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QPZOI  K/2 

Columbia  atotoetsitp 
intljeCitpoflrttigork 


(Eallwjr  of  ^yairtana  anb  Buv^ans 


Separimeni  of  ptjostoloiuj 
iPurrljajB?&  bg  ifjp 


•^  '  l*  rredericS.  L88, 

Oolnmbia  IMrvsity, 
HevYoric. 


Biochemical  Studies  of 
Sulfocyanates 


DISSERTATION 

SUBMITTED  IN    PARTIAL    FULFILMENT    OF  THE   REQUIRE- 
MENTS FOR  THE  DEGREE  OF  DOCTOR  OF  PHILOSOPHY 
IN  THE  FACULTY  OF  PURE  SCIENCE  OF 
COLUMBIA  UNIVERSITY  IN  THE 
CITY  OF  NEW  YORK. 


BY 

MAX  KAHN,  M.A.,  M.D. 

NEW  YORK  CITY 
1912 


Easton,  Pa.  : 
eschenbach  printing  company. 

1912. 


Biochemical  Studies  of 
Sulfocyanates 


DISSERTATION 

SUBMITTED  IN    PARTIAL    FULFILMENT    OF  THE  REQUIRE- 
MENTS FOR  THE  DEGREE  OF  DOCTOR  OF  PHILOSOPHY 
IN  THE  FACULTY  OF  PURE  SCIENCE  OF 
COLUMBIA  UNIVERSITY  IN  THE 
CITY  OF  NEW  YORK. 


BY 

MAX  KAHN,  M.A.,  M.D. 

NEW  YORK  CITY 
1912 


Easton,  Pa.: 

Eschenbach  Printing  Company. 

1912. 


Ql?20| 
K\2- 


TO  MY  MOTHER. 


CONTENTS. 


Tace. 

Acknowledgment  ..,. 6 

Chapter      I.  History 7 

Chapter    II.  The  ferric  chloride -ether  test 33 

Substances  that  give  a  similar  color  with  ferric  chloride 
or  that  may   interfere,    in   general,    with    the  sulfo- 

cyanate  test 39 

Conclusions 46 

Chapter  III.  Comparison  of  quantitative  methods 47 

Chapter  IV.  The  distribution  of  sulfocyanates  in  the  animal  body.  .  52 
Study  of  the  sulfocyanate  content  of  the  salivary  and 

other  glands  of  the  body 57 

Conclusions 58 

Chapter     V.  Metabolism  and  excretion  of  sulfocyanates 59 

Effect  of  administration  of  sulfur 60 

Effect  of  administration  of  sodium  sulfide 63 

Effect  of  administration  of  taurine 65 

Effect  of  administration  of  thiourea 67 

Effect  of  administration  of  acetonitrile 69 

Effect  of  administration  of  thioacetic  acid 69 

Effect  of  administration  of  alanin 70 

Effect  of  administration  of  glycocoll 71 

Effect  of  preventing  saliva  from  entering  the  stomach, 

upon  the  sulfocyanate  output 71 

Effect  of  fasting  upon  the  output  of    sulfocyanates.  .  72 

General  conclusions 73 

Chapter  VI.  Toxic  effects  of  potassium  sulfocyanate  on  animals  and 

plants 74 

On  frogs 74 

On  guinea  pigs 75 

On  dogs 76 

On  white  lupins 77 

On  timothy  grass  seed 78 

Bactericidal  properties  of  potassium  sulfocyanate ....  79 

Effect  of  KSCN  on  yeast   fermentation 80 

Effect  of  KSCN  on  the  souring  of  milk 80 

Conclusions 81 

Bibliography 82 

Biographical 84 

Publications 85 


ACKNOWLEDGMENT. 

Professor  William  J.  Gies  has  suggested  the  experiments 
embodied  in  this  dissertation;  he  has  aided  me  constantly 
in  my  work  with  his  cheering  encouragement  and  fruitful 
suggestions.  For  his  interest  in  my  labors  and  for  his  ma- 
terial aid,  I  wish  now  to  express  to  him  my  very  sincere 
gratitude.  By  his  designation  I  have  enjoyed  the  use  of  a 
grant,  for  the  purpose  of  this  research,  from  the  Dental  Society 
of  the  State  of  New  York. 

I  wish  to  thank  the  entire  staff  of  the  Department  of 
Biological  Chemistry  for  the  courtesy  and  cordiality  which 
they  have  always  shown  me. 

M.  K. 

Laboratory  of  Biological  Chemistry, 

College  op  Physicians  and  Surgeons, 

Columbia  University,  New  York, 

May  18,  1912. 


CHAPTER  I. 

HISTORY. 

The  slight  tha,t  Immanuel  Kant,  the  great  German  phil- 
osopher, had  cast  upon  chemistry  as  an  inexact  science  has 
long  ago  been  proven  to  be  undeserved;  for,  of  all  sciences, 
chemistry  (together  with  physics)  stands  preeminently 
forth  as  the  most  exact  and  most  mathematical  branch  of 
natural  philosophy.  Nevertheless,  if  one  were  to  take  a 
superficial  glance  over  the  literature  of  certain  topics  in  this 
science,  one  would  at  first  assuredly  become  impressed  with 
the  fact  that  uncertainty  is  the  fundament  of  this  study. 
Particularly  striking  is  the  history  of  the  experimentation 
and  theorizing  that  have  been  laboriously  undertaken  in 
the  study  of  sulfocyanate  in  the  animal  organism.  Denials, 
refutations,  contradictions  and  abnegations  can  be  found 
for  every  positive  statement  made  on  this  subject.  To 
every  Yea  there  is  an  emphatic  Nay,  which  is  in  turn  followed 
by  a  corroboration  and  again  by  doubt  and  interrogation. 
In  fact  over  the  whole  mass  of  literature  on  this  subject  one 
can  but  make  a  very  large  question  mark — and  sometimes 
several  exclamation  points  and  asterisks. 

One  is  sometimes  amazed  at  the  ridiculous  claims  and  un- 
founded deductions  that  certain  authors  make  for  the  sulfo- 
cyanates.  Ofttimes,  the  reader  would  fain  laugh  loudly 
at  the  absurd  statements  and  pedantic  phraseology  that  he 
meets  with  in  going  over  certain  papers  on  this  subject. 
I  confess  that  I  have  frequently  been  broadly  amused  at  the 
literary  antics  and  scientific  contortions  that  supposedly 
learned  men  deign  to  adopt  to  prove  what  they  consider 
an  important  point.  Like  the  schoolmen  of  old,  many  of  the 
contributors  to  this  subject  are  industriously  attempting  an 
answer  to  the  scientific  counterpart  of  that  most  "live" 
of  all  questions:  "How  many  angels  can  comfortably  dance 
on  the  point  of  a  needle."  However,  let  us  say  with  the  old 
Roman — Nihil  h  u m a n i  a  m e  alien u m  /> uto. 

The  history  of  saliva1  is  in  itself  a  very  interesting  chapter 
1  Hays:  Med.  News,  1904,  lxxxiv,  p.  582. 


8 

in  the  study  of  the  development  of  medicine.  In  the  days  of 
Hippocrates,  nothing  almost  was  known  of  physiology,  and 
the  salivary  secretions  were  thought  to  be  excrementitious. 
Galen  supposed  that  the  saliva  was  carried  to  the  mouth  by 
certain  veins.  Most  of  the  physicians  of  those  days  agreed, 
that  saliva  was  composed  of  water,  salt  and  spirit.  It  was 
used  as  a  medicament  in  certain  ailments.  We  are  told 
that  Vespasius,  when  visiting  Alexandria,  was  requested  by 
one  of  the  inhabitants  to  deign  and  expectorate  into  his  eye, 
in  order  to  be  relieved  of  blindness.  The  saliva  was  used 
as  a  balm  in  sores  and  pains  and  in  order  to  remove  the  pits 
of  small-pox.  The  athletes  of  ancient  Greece  used  to  soak 
their  plams  in  saliva  in  order  to  attain  grace  and  suppleness 
of  the  hands.  Even  now  when  a  husky  laborer  wants  to 
do  an  especially  hard  piece  of  work,  he  spits  into  his  palms, 
and  seems  to  derive  much  benefit  therefrom.  In  the  small 
villages  of  Russia  when  a  child  is  suffering  from  a  cutaneous 
disease  of  the  face,  it  is  taken  to  an  "old  woman"  who  mum- 
bles some  words  and  spits  several  times  into  the  mouth  of  the 
child — a  very  valuable  and  efficacious  remedy,  and  highly 
aesthetic ! 

Perhaps,  the  first  investigator  to  attempt  the  analysis 
of  saliva  scientifically  was  Hapel  de  la  Chenaye,1  who  in 
1780  published  his  results.  In  1790,  Berzelius,  the  great 
Swedish  chemist,  described  the  properties  of  the  enzyme 
ptyalin,  which  was  more  fully  studied  by  Leucho  in  1828. 

In  1 8 14,  Gottfried  Rheinhold  Treviranus2  reported  his 
results  of  the  study  of  the  chemistry  of  saliva.  "I  have 
found,"    he    writes,3    "in   the    saliva   two    substances    which 

doubtlessly  play  an  important  function The  second 

I  call  blood  acid  (Blutsaure) ,  which  is  found  also  in  the  blood 
and  which  has  the  following  characteristics :  With  a  saturated 
solution  of  iron  in  nitric  acid  or  in  dilute  sulfuric  acid,  it 
reacts  to  give  a  scarlet,  blood-like  coloration.  This  reaction 
is  also  obtained  upon  dropping  the  iron  solution  upon  the 

1  Chenaye:  Mem.  de  la  Soe.  Roy.  de  Medicine  de  Paris,  1780,  lxxxi,  p. 

325- 

2  Treviranus:  Biologie    oder    Physiologic    der    lebenden    Natur    fur 
Naturforscher  und  Aerzte,  1814,  iv,  p.  330. 

3  Treviranus:  My  translation. 


saliva.  If  alkalies  are  added  to  the  mixture  of  blood  acid 
with  iron,  an  orange-colored  precipitate  is  produced.  With 
silver  nitrate  a  black-brown  precipitate  forms.  Potassium 
cyanide  produces  no  effect.  Upon  this  acid  depends  the  color 
of  the  blood." 

The  empirical  name  of  "Blood-acid"  was  disliked  by  the 
chemists  of  those  days  who  desired  to  know  the  structure 
and  properties  of  this  substance.  When  Porret1  in  1820 
discovered  Rhodanwasserstoffsdure,  the  scientists  endeavored 
to  ascertain  whether  this  acid  was  present  in  the  animal 
organism.  Tiedemann  and  Gmelin2  in  1826,  confirmed 
lire's3  finding  that  the  "blood  acid"  in  the  saliva 
was  identical  with  the  sulfocyanic  acid  discovered 
by  Porret.  They  recommended  that  its  presence  should 
be  tested  for  in  all  the  secretions  and  excretions  of  the 
body.  They  were  not  sure  whether  urine  contained  it,  but 
strongly  urged  further  researches  along  that  line,  advising 
that  even  the  perspiration  be  examined  for  that  substance. 

Berzelius4  at  first  expressed  doubt,  but  later  gave  an 
unenthusiastic  confirmatory  report  as  to  its  presence  in  the 
saliva. 

Physiological  chemists  began  to  investigate  whether  the 
salts  of  hydrosulfocyanic  acid  are  localized  only  in  the  salivary 
secretions,  or  are  of  wide-spread  occurrence  in  the  animal 
body.  Researches  were  instituted  to  determine  the  absence 
or  presence  of  this  substance  in  the  secretions  of  the  lower 
animals.      Rough  quantitative  results  were  published. 

Mitscherlich5  had  the  opportunity  of  collecting  the  saliva 
from  the  parotid  gland  alone,  in  a  subject  who  had  an  ex- 
ternal fistulous  opening  of  that  gland.  He  reported  that  the 
amount  of  potassium  sulfocyanate  in  the  parotid  secretion 
was  three  parts  per  ten  thousand.  The  parotid  of  dog, 
horse  and  sheep,  he  affirmed,  also  contained  the  sulfocyanate. 

1  Porret:  Gilbert's  Ann.,  1820,  liii,  p.  184. 

1  Tiedemann  and  Gmelin:  Die  Verdauiing  nach  Versuchen,  1826,  i, 
p.  9. 

3  Ure:  Hays,  Loc.  cit. 

4  Berzelius:  Jahresbericht,  1827,  i,  p.  48. 

8  Mitscherlich:  Chcm.  Centralblatt,  1833,  p.  514. 


IO 

Oehl1  using  a  colorimetric  method  {i.  e.,  comparing  standard 
amounts  of  sulfocyanate  which  had  been  treated  with  ferric 
chloride,  with  the  saliva  which  had  been  similarly  treated) 
obtained  figures  almost  identical  with  those  of  Mitscherlich. 
Succeeding  observers  found  that  the  salt  under  discussion 
was  present  in  various  secretions  and  tissues.  Maly2  found 
traces  of  it  in  the  submaxillary  gland ;  Longet3  reported  positive 
results  in  the  sublingual  secretion .  Ellenberger  and  Hof  meister  * 
did  not  find  the  acid  in  the  tissues  of  the  horse,  cow,  sheep, 
hog,  goat.  Various  laboratory  workers  obtained  varying 
figures  for  the  quantity  of  sulfocyanate  in  the  mixed  human 
saliva.  Wright5  in  his  extensive  studies  on  the  physiology 
and  pathology  of  the  saliva  found  0.51-0.98  per  cent,  of  the 
salt.  In  1846,  Frerich8  stated  that  the  figures  of  the  English 
observer  were  too  high;  he  obtained  only  0.01  per  cent,  of 
potassium  sulfocyanate  in  the  saliva  of  a  healthy  person. 
Tillanus7  confirmed  the  findings  of  Frerich.  In  1850, 
Jacubowitsch8  carried  out  an  extensive  study  of  saliva. 
His  method  for  the  quantitative  determination  of  the  sulfo- 
cyanates  was  to  oxidize  the  sulfur  to  the  sulfate  ion  and  then 
to  precipitate  with  barium  and  weigh  the  precipitated  barium 
sulfate.  His  procedure  in  detail  was  as  follows:  He  took 
750  cc.  saliva,  and  extracted  several  times  with  alcohol. 
The  extract  was  filtered,  and  the  alcohol  was  driven  off  by 
evaporation  from  the  filtrate.  The  remaining  portion  was 
distilled  with  phosphoric  acid,  and  the  distillate  was  mixed 
with  barium  hydrate  and  barium  nitrate  and  heated  for  some 
time.      The  barium  sulfate  was  filtered  off  on  an  ashless  filter 

1  Oehl:  La    saliva  umana   studiata   colla    siringazione    dei    condotti 
gheandolari,  1864,  p.  177;  also  Constatt's  Jahresbericht,  1864. 

2  Maly:  Hermann's  Handb.  d.  Physiol.,  v,  p.  14. 

3  Longet:  Compt.  rend.  hebd.  des  seances  de  1'Academie  des  Sciences, 
1856,  p.  480. 

'  Ellenberger    and     Hofmeister:  Physiologie    der    Haussaugethiere, 
1890,  p.  495. 

*  Wright:  Lancet,  1841-42,  pp.  5,  72,  214,  262,  535,  563,  628,  672, 
813,  etc. 

6  Frerich:  Wagner's  Handworterbuch,  1846,  iii,  p.  766. 
'  Tillanus:  Constatt's  Jahresbericht,  1849,  p.  105. 

*  Jacubowitsch:  "De  saliva,"  Inaugural  Dissert.,  Dorpat,  1850. 


1 1 

paper,  dried,  ignited  and  weighed.  Jaeubovvitsch  found 
in  the  mixed  saliva  0.006  per  cent,  of  sulfoeyanate.  Like 
Mitscherlich,  he  found  this  substance  in  the  parotid  secretion 
of  man,  dog,  hocse  and  sheep.  Bidder  and  Schmidt1  corrob- 
orated the  findings  of  their  pupil  Jaeubovvitsch.  Yierodt2 
used  the  spectroscope  in  order  to  determine  the  amount  of 
potassium  sulfoeyanate  in  the  saliva.  He  found  in  one 
thousand  parts  of  the  secretion  0.16  parts  of  the  salt.  Leh- 
man3 obtained  0.0064-0.009  per  cent,  of  the  substance  under 
discussion  in  the  human  saliva.  On  the  other  hand, 
Tiegerstedt4  found  somewhat  higher  quantities  of  the  acid 
in  the  saliva — 0.1  part  per  one  thousand.  Bruylants,  an 
industrious  Belgian  scientist,  found  the  subject  of  sulfo- 
cyanates  in  the  saliva  in  a  very  vague  state.  His  researches 
will  be  discussed  later  (p.  18).  He  found  on  the  average  0.0374 
part  of  the  acid  in  a  liter  of  saliva. 

Immanuel  Munk6  in  1 869  began  a  study  of  the  sulfocyanates 
in  the  human  body.  His  method  for  the  quantitative  deter- 
mination of  the  acid  in  the  saliva,  I  shall  give  here  in  some 
detail:  The  saliva  was  filtered,  and  soda  was  added  to  the 
filtrate.  The  filtrate  was  evaporated  on  a  water  bath  to 
small  bulk  and  was  extracted  several  times  with  alcohol. 
The  extract  was  again  filtered  and  the  filtrate  was  evaporated 
to  dryness.  The  dry  residue  was  dissolved  in  some  water 
and  was  filtered.  The  filtrate  was  acidified  with  dilute  nitric 
acid,  and  enough  silver  nitrate  was  added  to  complete  pre- 
cipitation. The  precipitate  that  fell  out  was  composed  of 
silver  chloride  and  silver  sulfoeyanate.  This  was  collected 
on  an  ashless  filter  paper,  washed  with  water  carefully  and 
dried  at  ioo°  C.  It  was  then  fused  with  soda  and  sodium 
nitrate  in  a  silver  crucible.  The  excess  of  nitric  acid  was 
gotten  rid  of  by  adding  hydrochloric  acid  and  evaporating. 

1  Bidder  and   Schmidt:  Die   Verdauungssafte   und   der  Stoffweehsel, 
1852. 

aVierodt:  Die  Anwendbarkeit  des  Spektralapparates,   1873,  P-    J50. 
3  Lehman:  Zeitschr.  f.  Physiol.  Chem.,  1881,  v,  p.  302. 

*  Tiegerstedt:  Lehrbueh  der  Physiologie  des  Mensehen,  1891,  i,  p.  2  2  1 . 
5  Bruylants:   Bull,  de  l'Academie  de  Medicine  Bclgique,  1888,  p.  21. 

•  Munk:  Virehow's  Archiv.,  1869,  p.  350. 


12 

The  remainder  was  dissolved  in  water,  filtered,  and  the 
filtrate  precipitated  with  barium  chloride.  After  allowing 
the  precipitate  to  settle  for  twenty-four  hours,  the  barium 
sulfate  was  collected  on  an  ashless  filter  paper.  This  was 
dried,  ignited  and  weighed.  His  researches  were  confirmed 
in  1877  and  his  figures1  for  the  amount  of  sulfocyanates  in 
the  saliva  were  ten  to  forty  times  the  amount  that  Bruylants 
reported  in  1888.  Further  studies2  were  made  by  Munk 
and,  indeed,  he  is  the  most  reliable  and  scientific  investigator 
in  this  field. 

Before  proceeding  further  with  the  literature  on  the  analysis 
of  saliva,  it  is  advisable  to  pause  and  look  into  the  negative 
side  of  this  question. 

In  1843,  Blondlot3  criticized  the  findings  of  Tiedemann  and 
Gmelin.  I  shall  quote  him  verbally  in  his  own  language: 
"Que  penser  de  1' assertion  emisse  par  M  M.  Tiedemann  et 
Gmelin,  relativement  a  la  presence  dans  la  salive  de  l'acide 
hydrosulfocyanique?  D'abord  le  fait  sur  lequel  ces  auteurs 
s'appuient  pour  avancer  cette  opinion  etrange,  savoir,  la 
coloration  de  la  salive  en  rouge  par  le  perchlorure  de  fer, 
n'a  peutetre  reproduit  par  la  plupart  des  auteurs  qui  ont  voulu 
le  verifier,  et  je  puis  affirmer,  pour  mon  propre  compte,  que 
jamais  cette  experience  ne  m'a  reussi,  ce  qui  porte  a  croire 
que  les  savantes  professeurs  de  Heidelberg  ont  ete  induits 
en  erreur  par  quelques  circonstances  .  accidentelles  ou 
pathologiques.  Quant  au  procede  qu'ils  ont  mis  en  usage 
a  l'effet  d'isoler  ce  principe,  il  suffit,  pour  toute  critique, 
de  dire  quil  consiste  a  distiller  de  l'extrait  alcoholique  de 
salive  avec  de  l'acide  phosphorique  mediocrement  con- 
centre." 

Another  scientist  to  state  that  sulfocyanates  were  not  a 
normal  constituent  of  saliva,  was  Schiff.  He  advanced  the 
theory4  that  the  presence  of  the  rhodanate  is  only  a  test- 
tube  experience,  and  that  the  substance  does  not  occur 
in   the  living  body.     When  the  saliva,  he  said,   is  exposed 

1  Munk:  Deut.  Med.  Woch.,  1877,  p.  46. 

2  Munk:  Arch.  f.  d.  ges.  Physiol.  Bonn,  1895,  lxi,  p.  620. 

3  Blondlot:  Traite  Analytique  de  digestion,  1843,  p.  123. 

1  M.  Schiff:  Lecons  sur  la  physiologie  de  la  digestion,  1867,  i,  p.  147. 


*3 

to  the  air,  it  undergoes  spontaneous  decomposition.  The 
saliva  from  animals  was  collected,  and  put  into  a  number  of 
test  tubes.  He  tested  the  liquids  for  potassium  sulfocyanate 
at  twenty  minutes'  interval  of  exposure.  Those  which  were 
exposed  longer,  gave,  according  to  him,  very  much  higher 
results:  "En  effet,  chez  des  animaux  enrages,  l'experi- 
mentation  n'est  pas  facile,  on  ne  s'approche  pas  volontiers 
d'eux  pour  recueillir  leur  salive  au  fur  et  a  mesures  qu'elle 
se  forme,  mais  Ton  se  contents  de  leur  appliquer,  une  fois 
pour  toutes,  un  collecteur,  on  il  s'en  remasse  une  certaine 
quantite  que  Ton  examine  apres  quelque  temps." 

The  great  physiologist  Claude  Bernard  propounded  the 
theory  that  the  presence  of  sulfocyanates  in  the  saliva  was  a 
simple  result  of  tobacco  smoking,  and  that  those  that  did 
not  indulge  in  the  use  of  the  tobacco  leaves,  had  no  rhodanates 
in  their  body.  Later  on1  he  withdrew  his  absolute  denial, 
but  said  that  smokers  have  very  much  more  of  this  acid  and 
salts  in  the  saliva  than  non-smokers.  The  latter  statement 
was  taken  exception  to  by  very  many  observers;  preeminent 
among  whom  was  Hoppe-Seyler.2  He  flatly  denied  that 
smoking  had  any  effect  upon  the  amount  of  sulfocyanate 
excreted  in  the  saliva.  He  also  observed  that  the  dog's 
tissues  contained  no  traces  of  this  acid  or  its  salts,  thus  dis- 
proving such  authorities  as  Mitscherlich,  Gmelin  and  Jacubo- 
witsch.  But  lately,  Bunting3  put  the  very  pertinent  query 
whether  the  substance  that  gives  the  red  color  with  ferric 
chloride  is  really  potassium  sulfocyanate,  or  whether  it  is 
some  other  substance,  whose  structure  and  properties  have 
not  yet  been  described.  I  shall  discuss  this  author's  paper 
more  fully  in  a  later  chapter.      (p.  33). 

It  may  be  advisable  at  this  point  to  review  the  several 
attempts  that  have  been  made  to  affect  the  qualitative  deter- 
mination of  the  salts  of  sulfocyanic  acid.  The  characteristic 
test  that  was  discovered  by  Treviranus  was  for  many  years 

1  Bernard:  Lceons  sur  les  effcts  des  substances  toxiques  et  medi- 
camentenses,  1857,  p.  386. 

1  Hoppe-Seyler:  Lehrbuch  d.  Physiol.  Chem.,  1881,  p.  186. 
3  Bunting:  Dental  Cosmos,  1910,  Hi,  p.  1346. 


the  only  one  adopted  by  chemists  in  their  search  for  the 
thiocyanates.  In  fact,  with  the  discovery  of  Porret,  this 
acid  began  to  be  used  in  testing  for  very  slight  traces  of  iron. 
Several  other  reactions  were  subsequently  reported,  and  claims 
were  made  that  they  were  typical  only  of  the  rhodanates. 

The  ferric  chloride  test1  modified  somewhat  since  the 
days  of  Treviranus,  is  carried  out  as  follows:  To  a  little 
saliva  in  a  small  porcelain  crucible  or  dish,  add  a  few  drops 
of  dilute  ferric  chloride  and  acidify  slightly  with  hydro- 
chloric acid.  Red  ferric  thiocyanate  forms.  To  show  that 
the  red  coloration  is  not  due  to  iron  phosphate  add  a  drop  of 
mercuric  chloride,  when  a  colorless  mercuric  thiocyanate 
forms. 

In  1872,  Bottger2  recommended  the  adoption  of  the  fol- 
lowing test  as  corroborative  of  the  presence  of  sulfocyanates 
in  the  saliva.  He  took  strips  of  filter  paper  and  soaked 
them  in  a  tincture  of  guaiacum.  The  strips  were  then  dried 
and  drawn  through  a  1  :  2000  solution  of  copper  sulfate. 
When  a  drop  of  saliva  is  added  to  this  paper,  the  latter  be- 
comes blue,  if  the  thiocyanates  are  present.  This  test  seems 
to  have  been  neglected,  and  the  majority  of  modern 
physiological  chemistry  text-books  make  no  mention  of  it. 

Solera3  in  1877,  searching  for  a  new  test  for  the  rhodanates, 
made  the  following  discovery:  Thiocyanates  liberate  iodine 
when  acting  upon  iodic  acid.  He  prepared  a  test  paper 
according  to  these  directions:  Saturate  a  good  quality  of 
filter  paper  with  0.5  per  cent,  starch  paste  to  which  has  been 
added  sufficient  iodic  acid  to  make  a  1  per  cent,  solution  of 
iodic  acid,  and  allow  the  paper  to  dry  in  the  air.  Cut  it  in 
strips  of  suitable  size  and  preserve  for  use.  If  a  piece  of 
this  starch  paste-iodic  acid  paper  is  moistened  with  a  little 
saliva,  it  will  assume  a  blue  color  if  thiocyanates  are  present, 
due  to  the  liberation  of  iodine  and  the  subsequent  formation 
of  the  so-called  iodide  of  starch. 

Richard  Gscheidlen4  whose  labors  in  this  field  will  soon  be 

1  Hawk:  Practical  Physiological  Chemistry,  1910,  p.  56. 

*  Bottger:  Zcitschr.  f.  anal.  Chem.,  1872,  xi,  p.  350. 

5  Solera:  Maly's  Jahresber.,  1877,  vii,  p.  256. 

4  Gscheidlen:  Maly's  Jahresbd.  Tierchemie,  1874,  iv,  p.  91. 


15 

fully  considered,  adopted  the  Treviranus  test,  and,  three 
years  before  Solera,  also  prepared  a  thioeyanate  test  paper. 
Strips  of  filter  paper  are  soaked  in  a  mixture  of  ferric  chloride 
and  dilute  hydrochloric  acid,  and  dried  in  the  air.  Upon 
this  a  drop  of  saliva  is  let  fall  from  the  mouth.  The  paper 
will  become  red  if  the  test  is  positive. 

I  shall  briefly  describe  several  other  tests  for  the  identifica- 
tion of  sulfocyanates  in  the  saliva.  Colosanti1  found  that  if 
a  very  dilute  solution  of  thiocyanic  acid  or  its  potassium 
or  sodium  salt  is  treated  with  a  few  drops  of  copper  sulfate 
solution,  it  will  assume  a  beautiful  green  color.  He  cautioned, 
however,  that  before  applying  the  test  to  saliva,  the  latter 
must  be  freed  of  mucin  by  alcohol,  and  after  filtering,  the 
filtrate  should  be  concentrated  strongly  on  a  water  bath  and 
then  redissolved  in  a  little  water.  This  solution  is  then 
tested  as  above. 

In  the  same  year,  the  same  author2  following  out  his  in- 
vestigations of  1887-18883  reported  a  new  reaction  of  sulfo- 
cyanic  acid.  He  made  use  of  the  finding  of  Agostini4  on  the 
behavior  of  sugar  when  treated  with  gold  chloride. 
Colosanti's  discovery  hinges  upon  the  fact  that  when  a  very 
dilute  solution  of  thioeyanate  is  made  alkaline  with  sodium  or 
potassium  hydroxid,  it  will  give  a  very  pretty  violet  coloration 
if  treated  with  a  1  :  1000  solution  of  auric  chloride.  If  the 
sulfocyanate  is  not  very  dilute,  the  reaction  will  take  place 
in  the  cold,  otherwise  it  is  necessary  to  warm  the  reagents. 

In  1903,  Ganassini6  in  his  article  "  Complementa  al  mctodo 
Solera  e  nuovi  metodi  per  la  ricerca  delV  acido  sulj ocianico ," 
gave  several  new  reactions  for  the  rhodanates,  and  modified 
Solera's  test.  The  modification  consisted  in  this;  that  the 
author  in  analyzing  saliva  for  the  thiocyanates,  not  only 
determined  the  amount  of  iodine  liberated,  but  also  the 
quantity  of  cyanogen  iodide  that  resulted  according  to  the 

1  Colosanti:  Maly's  Jahresber.  d.  Tierchemie,  1890,  xix,  p.  72. 

2  Colosanti:  Maly's  Jahresber.  d.  Tierchemie,  1890,  xix,  p.  73. 

3  Colosanti:  Bull,  della  R.  Acad.  Medic,  di  Roma,  1887,  xiv,  p.  184. 

4  Agostini:  Ann.  di  chim.  e  farmac,  1886,  iv,  pp.  3,  228. 

8  Ganassini:  Biochcmisches  Centralblatt,  1904,  ii,  p.  361. 


i6 

equation: 

5KSCN  +  7HIO3  =  5KHS04  +  5CNI  +  I2  +  H20. 

The  several  new  tests  that  he  recommended  were :  (1) 
The  blue  coloration  that  thiocyanate  gives  with  ammonium 
molybdate  and  hydrogen  sulfide.  (2)  The  formation  of  lead 
sulfate  and  hydrocyanic  acid  upon  treatment  with  lead 
peroxide.     (3)  The  black  coloration  with  lead  tartrate. 

There  was  also  suggested  by  the  same  author  a  micro- 
chemical  method  for  the  determination  of  the  salts  of  sulfo- 
cyanic  acid.  With  mercurous  cyanide,  the  rhodanate  forms 
a  double  combination,  which,  upon  the  addition  of  iodic  acid, 
is  transformed  to  mercurous  iodide.  Polacci  in  his  report 
of  the  researches  from  the  chemical  and  toxicological  in- 
stitute of  Pavia  made  the  following  test  for  thiocyanates 
in  the  saliva:  To  some  pure  calomel  in  a  crucible  add  10-12 
drops  saliva,  and  stir  well.  Metallic  mercury  separates  if 
the  test  is  positive. 

Of  the  quantitative  analytical  methods  for  the  thiocyanates 
there  are  several  which  should  be  discussed,  some  briefly 
and  a  few  quite  extensively.  The  color  reaction  that  the 
rhodanates  give  with  the  chloride  of  iron  suggested  a 
colorimetric  scheme  for  the  quantitative  determination  of 
the  acid  and  its  salts.  Oehl1  in  1864  invented  a  color  scale 
for  the  measuring  of  the  amount  of  sulfocyanic  acid  in  the 
saliva.  Each  color  in  the  scale  was  compared  to  a  standard 
thiocyanate  solution  which  had  been  treated  with  iron 
chloride,  and  the  amount  noted  on  the  scale.  The  saliva, 
after  being  treated  according  to  Treviranus,  was  then  com- 
pared with  the  scale,  and  the  amount  found  and  recorded. 
Mitscherlich2  who  worked  almost  simultaneously  with  Oehl, 
and  also  used  a  colorimetric  method,  obtained  results  nearly 
identical  with  those  of  the  Italian  observer.  The  method  of 
Oehl  was  made  use  of  by  most  observers,  among  whom  can 
be  mentioned  Gscheidlen3  and  Pehl4  in  their  search  for  thio- 

1  Oehl:  La   saliva    umana   studiata    colla   siringazione   dei    condotti 
ghlandolari,  1864,  p.  177. 

2  Mitscherlich:  Loc.  cit. 

5  Gscheidlen:  Maly's  Jahr.  d.  Tierchemie,  1876,  vi,  p.  140. 
4  Pehl:  Maly's  Jahr.  d.  Tierchemie,  1877,  v">  P-  2°5- 


17 

cyanates  in  the  tissues  and  fluids  of  the  body. 

Vierodt1  in  1873  made  use  of  the  spectroscope  to  determine 
the  amount  of  thiocyanates  in  a  fluid.  About  twenty-five 
years  later,  Wroblewski2  in  his  analytical  study  of  saliva, 
also  came  to  the  conclusion  that  it  is  very  feasible  to  use 
the  spectroscope  for  the  determination  of  the  amount  of  sulfo- 
cyanate  in  the  salivary  secretions.  The  results  of  both  the 
above  observers  were  denied  by  Kriiss,3  who  stated  that  it  is 
quite  impracticable  to  apply  the  spectroscope  to  this  pur- 
pose. 

The  method  of  Colosanti4  also  lends  itself  to  colorimetric 
application,  similar  to  the  scheme  of  Oehl,  except  that  copper 
sulfate  is  used  instead  of  ferric  chloride. 

Albert5  went  one  step  further  in  this  colorimetric  process. 
He  caused  glasses  to  be  colored  various  shades  of  red,  by 
comparing  with  standard  thiocyanate  solutions  which  had 
been  treated  with  dilute  ferric  chloride.  He  then  applied 
a  Fleischman  hemoglobinometer  to  his  purpose,  and  thus 
had  a  very  efficient  method,  as  he  states,  for  the  quantitative 
analysis  of  hydrosulfocyanic  acid  or  its  salts. 

The  Dental  Society  of  America  recommended  the  following 
test  :8 

"Take  1  cc.  of  the  specimen  in  tube  A,  and  1  cc.  of  1  :  2000 
NH4SCN  in  tube  B.  Add  two  drops  of  a  5  per  cent,  solution 
of  ferric  chloride  to  each  tube;  add  water  to  tube  B  until 
its  color  matches  that  of  the  specimen.  Read  the  scale  in 
thousandths  and  ten  thousandths.  Care  must  be  taken  to 
have  the  bottom  of  the  meniscus  on  the  line.  If  these  tubes 
are  introduced  in  the  colorimeter,  the  readings  can  be  made 
more  accurately.  If  later,  diacetic  acid  is  found,  a  correction 
is  made  in  the  finding." 

The    gravimetric    methods    have    quite    a    short    history. 

1  Vierodt:  Loc.  cit. 

1  Wroblewski:  Chem.  Zentralblatt,  1897,  ii,  p.  532. 

3  Kriiss:  Maly's  Jahresb.  d.  Tierchemie,  1897,  xx\'ii,  p.  368. 

4  Colosanti:  Loc.  eit. 

5  Albert:  Lancet,  1898,  i,  p.  494. 

•  Ferris  and  Schiadieck:  Dental  Cosmos,  191 1,  liii,  p.  1297. 


i8 

Jacubowitsch1  in  his  inaugural  dissertation  employed  a 
gravimetric  process  which  has  already  been  described. 
Munk2  also  used  a  similar  procedure. 

Bruylants3  wanted  to  prove  positively  the  presence  of 
sulfocyanic  acid  in  the  saliva.  He  seemed  to  have  strong 
doubts  on  this  question.  He  carried  out  the  following  ex- 
periments: He  collected  iooo  cc.  of  saliva  and  added  some 
chloroform  as  a  preservative  and  then  he  made  it  alkaline 
in  reaction.  After  evaporating  to  200  cc,  he  added  25  cc. 
hydrochloric  acid  and  then  extracted  several  times  with 
ether.  The  collected  ether  extract  was  shaken  with  15  cc. 
water  and  a  little  ferric  chloride — -enough  to  give  the  maximum 
red  color  to  the  water  layer,  i.  e.,  upon  further  adding  the 
iron  halide  no  increase  in  color  was  to  be  noticed.  The 
red,  aqueous  layer  was  acted  upon  by  ammonia  until  it  be- 
came completely  decolorized;  it  was  then  warmed  and  filtered, 
and  the  nitrate  divided  into  two  parts. 

Part  one  was  evaporated  to  dryness  and  dissolved  in  ab- 
solute alcohol;  the  alcohol  was  then  driven  off,  and  the  re- 
mainder dissolved  in  water  and  precipitated  with  lead  acetate. 
After  allowing  to  stand  for  twenty-four  hours,  the  crystalline 
precipitate  was  filtered  off  and  dried  at  105 °  C.  and  weighed. 

Part  two  was  evaporated  to  10  cc.  and  distilled  with  2  cc. 
concentrated  hydrochloric  acid.  The  distillate  was  collected 
under  water  and  was  treated  with  zinc  and  sulfuric  acid  at 
30 °  C.  In  the  gases  produced  he  found,  hydrocyanic  acid, 
hydrogen  sulfid  and  methylamin. 

Bruylants  also  advocated  a  rapid  though  rough  method 
for  the  cyanate  determination :  A  portion  of  the  saliva 
was  treated  with  a  few  drops  of  chloroform,  and  filtered. 
To  about  10-15  cc.  of  the  clear  filtrate,  he  added  one  to  two 
drops  of  concentrated  hydrochloric  acid  and  an  equal  number 
of  drops  of  ferric  chloride  solution.  After  several  hours' 
standing,  he  compared  the  color  with  a  standard  solution  of 
ammonium  sulfocyanate,  according  to  the  method  of  Pehl.4 

1  Jacubowitsch :  Loc.  cit. 

2  Munk:  Loc.  cit. 

3  Bruylants:  Maly's  Jahresbericht  d.  Tierchemie,  1888,  xviii,  p.  134. 
*  Pehl :  Loc.  cit. 


19 

The  Belgian  scientist  was  very  emphatic  in  denying  that  the 
rhodanates  were  typical  of  the  saliva,  since  he  contended 
that  they  are  to  be  universally  found  in  the  animal  tissues. 
As  has  been  said  before,  however,  his  figures  for  the  thio- 
cyanate  content  of  the  saliva,  were  very  markedly  lower 
than  those  of  Munk  or  Gscheidlen. 

I  might  here  just  mention  the  method  of  Volhard'  which 
he  reported  in  his  paper  entitled  "  Die  Anwendung  des 
Sehwefelcyanammonium  in  der  Massanalyse." 

Perhaps,  the  most  exact  of  the  methods  for  the  quantitative 
determination  of  the  thiocyanates  is  the  iodometric  titrating 
process.  Rupp  and  Schied2  and  later  Rupp3  alone  wrote 
on  this  method.  The  underlying  principle  of  this  method 
is  the  fact  that  sulfocyanate  solutions,  treated  with  sodium 
bicarbonate,  decolorize  large  amounts  of  iodine,  cyanogen 
iodide  being  formed,  according  to  the  following  equation: 

CNSK  +  4I2  +  4H20  -  H2S04  +  6HI   +  KI   +  CNI. 

The  process  is  finished  in  four  hours  at  ordinary  tempera- 
ture. Upon  carefully  acidifying  with  hydrochloric  acid, 
the  potassium  iodide  is  changed  to  potassium  chloride  and 
hydriodic  acid  is  formed;  the  latter  acts  on  the  cyanogen 
iodide  to  form  hydrocyanic  acid.  The  whole  process  can  be 
expressed  thus: 

CNSK  +  31,  +  4H20  -  H2S04  +  5HI  +  KI  +  HCN 

that  is,  one  molecule  of  sulfocyanic  acid  is  equivalent  to  six 
molecules  of  iodine. 

I  shall  briefly  describe  the  procedure  for  the  analysis  of 
sulfocyanates  according  to  this  method.4  The  following 
reagents  are  necessary:  (a)  Dilute  nitric  acid  1  :  100;  (b) 
silver  nitrate  3  per  cent. ;  (c)  clean  infusorial  earth  which  has 
been  washed  in  acid;  (d)  sodium  bicarbonate,  chemically  pure; 
(e)  potassium  iodide;  (/)  N  1/10  iodine  solution;  (g)  N  1/10 

1  Volhard:  Liebig's  Ann.  d.  Chem.  u.  Pharm.,  1883,  cxc,  p.  24. 
a  Rupp  and  Schied:  Bcr.  deut.  chem.  Ges.,  1902,  xxxv,  p.  2 191. 

3  Rupp:  Arch.    d.    Pharm.,    1905,    ccxxxxiii,    p.    358;   also  in  Chem. 
Central.,  1905,  ii,  p.  1228. 

4  Neuberg:  "Der  Ham,"  1911,  i,  p.  542. 


20 

sodium  thiosulfate  solution;  (h)  hydrochloric  acid  solution 
10  per  cent.;  (t)  starch  solution  2  per  cent. 

The  analysis  is  conducted  in  the  following  way:  The 
fluid  to  be  examined  is  filtered,  and  the  filtrate  heated  to 
throw  down  any  albumin,  etc.,  and  again  filtered.  Acidify 
with  very  dilute  nitric  acid  and  add  an  excess  of  silver  nitrate. 
In  order  to  cause  complete  separation  add  some  infusorial 
earth  and  stir  and  warm  ten  minutes.  Add  a  little  more 
silver  nitrate  to  see  that  precipitation  is  complete.  Filter 
under  diminished  pressure  on  a  filter  paper  stuck  in  a 
platinum  cone.  Make  sure  that  the  filtrate  is  clear.  Wash 
several  times  with  nitric  acid.  Transfer  to  a  wide-necked 
glass  container  with  water,  and  add  3  grams  sodium  bi- 
carbonate to  alkaline  reaction.  Add  3  grams  potassium 
iodide  and  shake  slightly  until  the  solution  is  clear.  Add 
N  1  / 10  iodine  solution  until  it  assumes  a  brown  color.  Shake 
slightly  and  let  stand  in  a  dark  place  for  four  hours.  The 
solution  is  carefully  acidified  with  10  per  cent,  hydrochloric 
acid.  Add  a  few  cubic  centimeters  of  the  starch  solution. 
Titrate  with  A7  1/10  sodium  thiosulfate  until  a  lemon-yellow 
color  is  obtained. 

Kabdebo1,  a  Hungarian  observer,  in  his  study  of  the  origin 
and  fate  of  the  sulfocyanates  in  the  body,  used  the  iodometric 
method  of  Rupp  and  Schied  in  his  analyses. 

Hydrosulfocyanic  acid  was  at  first  supposed  to  be  typical 
of  the  salivary  secretion,  and  it  was  claimed  that  this  acid 
is  not  found  in  the  other  fluids  of  the  body.  However,  later  on, 
observers  found  the  acid  in  almost  every  organ  and  fluid 
of  the  body.  Funke2  attempted  unsuccessfully  to  determine 
the  origin  of  the  thiocyanates  in  the  human  body.  The 
only  conclusion  that  he  came  to  was  that  the  rhodanates 
were  a  constant  component  of  the  salivary  secretions. 
Carpenter3  states  that  the  substance  is  absent  in  the  saliva  and 
urine  of  herbivora.  Treviranus  suggested  that  it  is  present  in 
the  blood,  and  Tiedemann  and    Gmelin  remarked   that   it  is 

1  Kabdebo:  Jahresber.  der  Tierchem.,  1907,  xxxvii,  p.  401. 

2  Funke:  Lehrb.  d.  Physiol.,  1858,  i,  p.  220. 

3  Carpenter:  Human  Physiology,  1848,  p.  134. 


21 

present  in  most  of  the  secretions  of  the  body.  Gscheidlen1 
found  it  in  the  urine  of  most  subjects.  Lea2  says  that  it  occurs 
only  in  the  urine  of  those  animals  which  excrete  their  nitrogen 
chiefly  as  urea.  In  1 869,  Leared3  found  it  in  the  blood.  In  the 
first  months  of  life  of  human  beings,  Pribram4  did  not  find  any 
thiocyanates.  Nencki8  examined  the  gastric  juice  of  a  dog 
for  this  acid  and  found  0.005  gram  per  liter  of  the  secretion. 
He  made  sure  that  the  gastric  juice  was  free  from  saliva, 
or  else  his  results  would  not  have  been  of  any  scientific  value. 
In  1900,  Muck5  reported  that  he  had  discovered  traces  of  the 
rhodanates  in  the  nasal  and  conjunctival  secretions.  Musso7 
in  his  researches  on  sulfur  in  milk,  discovered  that  traces  of 
hydrosulfocyanic  acid  was  also  present  in  this  fluid.  This 
was  corroborated  by  Bruylants8  who  reported  that  he  found 
0.0016  gram  per  1000  grams  of  milk.  The  latter  observer 
also  analyzed  ox-gall,  and  gave  the  following  figure,  0.01 
gram  per  1000.  In  the  cystic  fluid  of  the  abdomen  he  found 
0.0007  gram  per  1000;  in  the  fluid  of  a  hydrocele  0.00055 
gram  per  1000.  He  did  not  find  this  substance  in  the  urine 
of  individuals  suffering  from  podagra. 

Various  and  conflicting  results  have  been  reported  as  to 
the  presence  of  the  thiocyanates  in  the  fluids  of  the  lower 
animals.  As  was  stated  before,  Mitscherlich,  Gmelin  and 
Jacubowitsch  found  it  in  the  saliva  of  man,  dog,  horse  and 
sheep.  Ellenberger  and  Hofmeister  did  not  find  it  in  the 
secretions  of  horse,  cow,  sheep,  hog,  or  goat.  De  Souza9  found 
it  in  the  blood,  saliva,  pancreatic  juice  and  bile  of  human 
beings.  Bruylants  stated  that  the  thiocyanates  were  absent 
in  reptiles  or  fowls.     Hoppe-Seyler  also  found  none  of  this 

1  Gscheidlen:  Loe.  cit. 
a  Lea:  Chemical  Basis  of  the  Animal  Body. 
3  Leared:  Proc.  Roy.  Soc.  London,  1869,  xviii,  p.  16. 
1  Pribram:  Jahrb.   d.    Physiol,   u.    Pathol,   d.  ersten   Kindes  alters, 
1868,  i,  p.  148. 

s  Nencki:  Berichte  der  deut.  chcm.  Ges.,  1895,  xxviii,  p.  1318. 

•  Muck:  Munch,  mediz.  Wochenschr.,  1900,  xlvii,  p.  1168. 

7  Musso:  Berich.  f.  Physiol.  Chem.,  1877,  vii,  p.  168. 

8  Bruylants:  Loe.  cit. 

•  De  Souza:  Journal  Physiology,  1907,  xxxv,  p.  332. 


22 

substance  in  dogs.  In  a  calf,  twenty-two  days  old,  Bayer1 
found  the  rhodanates. 

The  presence  of  the  salts  of  hydrosulfocyanic  acid  in  the 
living  beings,  gave  rise  to  speculations  as  to  its  origin  and 
function  and  fate.  We  will  neglect  here  the  suggestion 
made  by  Claude  Bernard  that  the  rhodanates  are  really  a 
foreign  substance  introduced  into  the  human  body  through 
the  use  of  tobacco,  and  the  other  theory  advanced  by  Schiff 
that  the  thiocyanates  are  really  absent  in  the  saliva,  but 
are  produced  there  on  exposure,  due  to  spontaneous 
decomposition. 

Richard  Gscheidlen2  demonstrated  to  his  own  satisfaction 
that  the  thiocyanates  are  first  produced  in  the  saliva,  and 
then  it  passes  into  the  stomach,  and  via  the  blood,  to  all 
tissues  of  the  body.  He  did  the  following  researches  upon 
living  animals:  He  caused  the  ducts  of  the  salivary  glands 
to  empty  themselves  outside  of  the  mouth.  He  then  daily 
collected  the  saliva  and  tested  for  the  thiocyanates,  and  al- 
ways obtained  positive  results.  When,  however,  he  ex- 
amined the  blood  of  the  animal  or  its  urine,  he  always  found 
negative  results.  This  proved  to  him  conclusively  that  the 
thiocyanates  are  first  formed  in  the  salivary  glands.  His 
attempts  to  isolate  sulfocyanates  from  the  urine,  were  suc- 
cessful according  to  his  report,  only  when  the  dogs  swallowed 
their  saliva.  But  Thudichum3  wrote  in  1877  that  he 
could  not  duplicate  the  results  of  Gscheidlen,  even 
though  he  followed  his  methods.  The  latter,  never- 
theless, reported  that  in  spite  of  Thudichum's  findings,  he 
could  still  isolate  the  KSCN.  The  paper  in  which  Gscheidlen 
refutes  the  English  observer  is  entitled:  "  Wiederlegung  der 
von  Hern  Thudichum  erhobenen  Einwande." 

The  work  of  Gscheidlen4  would  find  some  corroboration 
in  the  studies  of  Longet5  a  summary  of  whose  findings,  I 

1  Bayer:  Maly's  Jahresber.  d.  Tierchemie,  1876,  vi,  p.  172. 

2  Gscheidlen:  Pfluger's  Arch.,  1877,  xiv,  p.  401. 
'Thudichum:  Maly's  Jahr.  d.  Tierchemie,  1877,  vii,  p.  205. 
*  Gscheidlen:  Loc.  cit. 

8  Longet:  Compt.  rend.  hebd.  des  seances  de  l'Academie  des  sciences, 
1856,  p.  480. 


23 

shall  give  here:  (a)  Potassium  sulfocyanate  is  a  normal 
and  constant  component  of  saliva.  (6)  It  is  present  not 
only  in  the  mixed  saliva,  but  also  in  the  secretions  of  the 
parotid,  submaxillary,  and  sublingual  individually,  (c) 
The  presence  of  potassium  thiocyanate  is  characteristic  of  the 
saliva,  because  it  is  not  found  in  the  other  animal  secretions, 
(d)  The  amount  of  thiocyanate  is  only  proportional  to  the 
concentration  of  saliva. 

In  1901,  Grober1  refuted  the  findings  of  Schiff,2  agreed 
somewhat  with  Longet,  and  advanced  another  theory  as  to 
the  origin  of  the  substance  under  discussion.  He  also  found 
that  in  human  beings,  KSCN  is  only  present  in  the  saliva. 
Unlike  Schiff,  he  not  only  did  not  find  an  increase  in  the  thio- 
cyanate content  of  saliva  upon  exposure,  but  in  fact,  he  noticed 
that  the  amount  of  KSCN  became  less  with  the  duration  of 
its  separation.  He  was  emphatic  in  denying  that  it  had 
anything  to  do  with  smoking  tobacco  or  using  nicotine. 
He  noticed  an  increase,  however,  when  prussic  acid  was  taken 
in  very  minute  doses  by  his  subjects.  The  excretion  of 
KSCN,  he  suggested,  depended  only  on  the  accumulation 
and  disintegration  of  proteids  in  the  body. 

Fen  wick,3  in  a  laborious  attempt  to  discover  whether  the 
sulfocyanates  had  any  specific  relation  to  various  diseases, 
did  not  come  to  any  conclusions  on  this  point,  as  some  later 
writers  have  done  on  this  continent.  He,  however,  found 
that  there  exists  a  relationship  between  the  bile  and  the 
KSCN  in  the  body.  He  found  that  the  sulfur  in  the  thio- 
cyanate was  derived  from  some  sulfur  compound  of  the  bile. 
When  he  diverted  the  bile  from  the  alimentary  canal,  he 
failed  to  discover  any  rhodanate  in  the  saliva. 

De  Souza4  negated  the  results  of  Gscheidlen.  He  found 
on  the  contrary,  that  instead  of  the  KSCN  passing  from  the 
saliva  into  the  blood,  it  travelled  in  the  other  direction.  He 
stated  that  the  sulfocyanates  pass  out  from  the  blood  into 

1  Grober:  Deut.  Arch.  f.  clin.  Med.,  1901,  lxix,  p.  243. 
3  Schiff:  Loe.  cit. 

3  Fenwick:  Medic.  Chirur.  Transactions,  1882,  Ixi,  p.  118. 
*  De  Souza:  Loc.  cit. 


24 

the  saliva,  pancreatic  juice,  bile  and  urine.  In  the  sub- 
maxillary saliva,  pancreatic  juice  and  bile  the  concentration 
is  always  less  than,  and  appears  to  depend  upon,  the  con- 
centration in  the  blood.  The  concentration  in  the  urine, 
on  the  other  hand,  he  said,  may  be  greater  or  less  than  the 
concentration  in  the  blood,  and  is  diminished  by  sodium 
sulfate  diuresis.  He  also  noticed  that  sulfocyanates  in  the 
food  are  readily  absorbed  and  remain  as  such  in  the  body 
for  a  considerable  time.  After  injection  or  feeding,  the 
parotid  saliva  contains  less  sulfocyanate  than  the  sub- 
maxillary, and  still  less  than  the  blood  serum. 

Brubaker1  seems  to  agree  with  Gscheidlen,  but  is  more 
certain  than  the  latter  author,  for  he  states  in  his  text  book 
that  the  sulfocyanates  in  the  body  were  derived  from  the 
secretions  of  the  parotid  glands  only. 

Some  authors  have  thought  it  possible  that  the  rhodanates 
in  the  saliva  were  a  kind  of  vicarious  excretion,  similar  to 
the  excretion  of  urea  in  saliva  in  cases  of  nephritis.2 

What  is  the  chemical  origin  of  the  sulfocyanates  in  the 
living  body?  This  indeed  is  a  question  the  answer  to  which 
it  is  very  difficult  to  give.  Theories  galore  have  been  ad- 
vanced and  objected  to  on  this  question.  Funke  in  18583 
gave  up  in  despair  the  attempt  to  solve  this  problem. 
Florian4  essayed,  but  also  was  unable  to  give  a  satisfactory 
answer.  The  only  thing  that  he  became  convinced  of  finally 
was  that  the  thiocyanates  are  a  constant  and  normal  com- 
ponent of  the  saliva.  Bruylants5  endeavored  to  prepare  the 
sulfocyanic  acid  from  proteids.  He  took  egg  albumin  and 
serum  albumin  as  his  starting  materials.  His  researches 
were  crowned  with  success.  The  procedures  that  he  followed 
were  three  in  number:  (a)  Dry  distillation  of  the  albumin 
gave  him  a  yield  of  0.205-0.224  per  cent.  (6)  Fusion  with 
potassium  hydroxide  or  (c)  boiling  with  the  same  hydrate, 
gave    somewhat   better   results.     It   must   be   noticed   here 

1  Brubaker:  "Text  Book  of  Physiology,"  1908,  p.  156. 

2  Leube:  Deutsche  Arch.  f.  Clin.  Med.,  1904,  lxvi,  p.  80. 

3  Funke:  Lehrbuch  d.  Physiol.,  1858,  p.  220. 

*  Florian:  Gaz.  medicalle  de  Paris,  1884,  p.  354  and  1889,  p.  317. 
'■  Bruylants :  Loc.  cit. 


25 

that  these  were  test  tube  experiments,  and  not  at  all  to  be 
taken  as  conclusive  evidence  that  similar  processes  are  at 
work  in  the  quick  body.  As  one  of  my  old  professors  used 
to  say:  "Two  and  two  in  the  laboratory  are  not  the  same 
two  and  two  in  the  living  being." 

Studies  were  undertaken  by  various  chemists1  to  determine 
the  fate  of  certain  nitriles  when  fed  to  animals.  Aceto-, 
propio-,  butyro-,  and  capro-nitriles  were  especially  investi- 
gated, with  the  hope  of  finding  that  they  may  be  the  cause 
of  the  origin  of  certain  substances  as  the  thiocyanates ;  but 
the  results  were  unsatisfactory.  The  toxic  effects  of  these 
compounds  will  be  spoken  of  in  due  time. 

Kossel2  found  that  adenin  yielded  hydrocyanic  acid  upon 
heating.  Gautier3  reported  that  upon  hydration  of  xanthin, 
prussic  acid  was  formed.  These  attempts  and  successes 
for  the  cyanide  give  us  hope  that  we  will  soon  discover  the 
forerunner  of  the  sulfocyanate.  Martinotti4  reviews  some- 
what superficially  the  literature  on  this  subject.  Willianen,5 
a  Russian  scientist,  found  that  the  urine  of  rabbits,  which 
ordinarily  contains  no  trace  of  rhodanates,  will  give  a  positive 
reaction  for  that  substance  if  the  animals  are  fed  glycocoll, 
creatinin  and  adenin  (weak).  It  also  seems  apparent  to  the 
author  that  the  amino  acids  as  well  as  the  other  named 
substances,  which  on  decomposition  give  hydrocyanic  acid, 
are  also  the  source  of  the  thiocyanates  in  the  living  organism. 

Upon  oxidation  of  albumins,  Plimmer6  produced  hydro- 
cyanic acid. 

G.  Kabdebo7  in  his  studies  in  this  field,  discovered  that 
the  administration  of  acetonitrile  lessened  the  oxidation  of 
sulfur  in  the  body.     He  also  reported  that  the  neutral  sulfur 

'Lang:  Arch.  f.  exper.  Path.  u.  Pharm.,  1894,  xxxiv,  p.  247;  and 
Giacosa:  Zeit.  f.  Physiol.  Chem.,  1883,  viii,  p.  95. 

2  Kossel:  Berliner  Klin.  Woch.,  1889,  No.  19. 

3  Gautier:  Chimic  biologique,  Paris,  1892,  p.  230. 

4  Martinotti:  Central  f..  Bakteriol.  u.  Parasitkunde,  1896,  xix,  p.  142. 
8  Willianen:  Biochem.  Centrall.,  1906,  v,  p.  477. 

*  Plimmer:  Jour,   of  Physiology,    1903-1904,   xxxi,   p.   65  and   1904, 
xxxii,  p.  51. 

7  Kabdebo:  Jahresber.  der  Tierehemie,  1907,  xxxvii,  p.  401. 


26 

of  the  body  unites  with  the  acetonitrile  to  form  hydro- 
sulfocyanic  acid.  De  Souza  in  his  observations1  on  the 
elimination  of  the  sulfocyanates  from  the  blood,  and  their 
supposed  formation  in  the  salivary  glands,  found  that  upon 
feeding  the  acetonitrile  to  dogs  (which  according  to  him  have 
no  rhodanates  in  their  fluids) ,  sulfocyanates  were  found  not 
only  in  the  urine  but  also  in  the  saliva  and  blood  serum. 

The  question,  however,  to  my  mind,  is  far  from  settled. 

It  is  interesting  at  this  point  to  know  what  normal  or 
pathologic  conditions  effect  the  secretion  of  KSCN  in  the  body 
in  general  and  in  the  saliva,  in  particular.  As  Kriiger2 
remarks  there  are  three  theories  concerning  the  thiocyanates : 

i .  That  KSCN  is  absent  altogether. 

2.  That  KSCN  is  present,  but  that  it  is  a  product  of  de- 
composition of  saliva. 

3.  That  KSCN  is  a  normal  constituent  of  the  saliva. 
Supposing  that  the  third  theory  is  accepted,  what  causes 

an  increase  in  the  output  of  KSCN  in  the  saliva? 

Claude  Bernard,3  as  has  been  stated  before,  reported  that 
smoking  tobacco  causes  an  increase  in  the  amount  of  thio- 
cyanic  acid  in  the  saliva.  This  was  contradicted  by  Hoppe- 
Seyler3  who  stated  that  smoking  had  no  effect  at  all  on  the 
rhodanates  content  of  the  saliva.  Gscheidlen3  found  the 
urine  of  smokers  richer  in  KSCN  than  the  urine  of  non- 
smokers.  Grober3  stated  that  the  amount  of  thiocyanates 
in  the  saliva  does  not  depend  upon  smoking  or  using  nicotine 
in  any  form.  Nevertheless,  Kriiger3  is  quite  positive  that 
tobacco  smoking  increases  the  amount  of  KSCN  in  the  saliva 
two  to  three  times.  Dr.  Lothrop  tells  me  that  in  his  studies 
with  Professor  Gies4  he  did  not  notice  any  variation  in  the 
reaction  for  sulfocyanates  in  smokers  and  non-smokers. 

Experimentally,  KSCN  can  be  diminished  in  the  saliva 
by  prolonged   stimulation  of  the  salivary  glands.5     In  his 

1  De  Souza:  Loc.  cit. 

2  Kriiger:  Zeitsch.  f.  Biologie,  1899,  xxxvii,  p.  6. 

3  Loc.  cit. 

4  Lothrop  and  Gies:  Journal  Allied  Dental  Soc,  1910,  v,  p.  4;  1911, 
vi,  p.  65. 

6  Schneider:  Am.  Jour.  Physiol.,  1901,-  v,  p.  274. 


27 

experiments,  Schneider  uniformly  found  that  the  parotid 
saliva  is  always  richer  in  KSCN  than  the  submaxillary 
secretion,  collected  from  the  same  individual  at  the  same 
time.  Grober1  found  that  feeding  of  hydrocyanic  acid  causes 
an  increase. 

Many  other  circumstances  have  been  accredited  as  the 
influencing  factors  of  the  presence  or  absence  of  the  thio- 
cyanates  in  the  saliva.  The  variety  of  food,  according  to 
Bruylants,1  has  no  effect  upon  the  thiocyanates,  which  is 
formed  in  the  organism. 

Is  the  condition  of  the  teeth  a  causative  factor  for  the  in- 
crease or  decrease  of  thiocyanates?  Longet1  investigated 
this  question  and  came  to  the  conclusion  that  the  sick  or 
healthy  condition  of  the  teeth  has  nothing  at  all  to  do  with 
the  presence  or  absence  or  amount  of  the  potassium  thio- 
cyanate  in  the  saliva.  In  1899,  Kriiger1  gave  a  very  emphatic 
affirmation  of  the  finding  of  Longet.  Lately,  however, 
Low2  and  Waugh3  found  that  most  patients  who  have  caries 
of  the  teeth  have  no  thiocyanates  in  their  saliva.  Lothrop 
and  Gies4  were  unable  to  corroborate  these  findings. 

Various  diseases  that  attack  the  organism  in  general,  are 
said  to  cause  changes  in  the  potassium  sulfocyanate  output 
in  the  saliva.  Some  pseudo-scientists  have  gone  so  far  as 
to  say  that  by  analyzing  the  saliva  in  various  illnesses  one 
could  make  diagnoses  of  those  affections :  Fenwick5  analyzed 
the  saliva  for  KSCN  in  numerous  disturbances  of  health,  but 
could  come  to  no  conclusion,  except  that  he  found  variations 
in  the  amount  of  thiocyanates  in  the  saliva.  Kyle8  advocates 
the  examination  of  saliva  (sialosemiology)  in  various  diseases, 
as  for  example  hay  fever,  and  assures  us  that  positive 
inferences  can  be  drawn  from  the  analyses.  In  1908,  there 
appeared    a   very    comical    article7   on   this    so-called    sialo- 

1  Loc.  cit. 

a  Low:  Dental  Cosmos,  191 1,  liii,  p.  1269. 

3  Waugh:  Dental  Cosmos,  1910,  Hi,  p.  170  and  420. 

*  Lothrop  and  Gies:  Loc.  cit. 
8  Fenwick:  Loc.  cit. 

•  Kyle:  Jour.  Am.  Med.  Assoc,  1907,  xlix,  p.  402. 
'  Lcroy:  N.  Y.  Med.  Jour.,  1908,  lxxxxvii,  p.  448. 


28 

semiology.  The  author  makes  the  following  "humorous" 
statements:  Absence  of  potassium  thiocyanate  indicates 
various  nervous  lesions,  as  for  example,  epilepsy,  paralysis 
and  dementia  praecox( !) .  The  reappearance  of  sulfocyanate 
in  typhoid  fever  is  hailed  as  a  very  evident  sign  of  the  return 
of  good  health.  Excess  of  this  marvelous  chemical  indicates 
(sic)  heart,  brain  or  kidney  lesions.  Boding  good  to  no 
one  is  the  phenomenon  of  the  following  misbehavior  of  the 
rhodanate:  In  cases  where  the  reaction  for  sulfocyanate 
gives  a  dark  brown  color  instead  of  a  brick-red  color,  and  if 
this  reaction  is  noticed  time  and  again  in  the  same  patient, 
you  may  notify  the  friends  of  a  fatal  issue(!?). 

The  great  Russian  physiologist,  Pavlov,1  studied  the 
effect  of  psychic  stimulation  upon  the  secretion  of  the  various 
fluids  of  the  body.  His  brilliant  researches  which  were 
crowned  with  remarkable  success  do  not  form  a  part  of  this 
paper.  But  it  is  in  this  connection  that  one  may  mention 
the  finding  of  Eberle,2  that  in  states  of  excitement,  worry  or 
anger  there  is  a  marked  increase  in  the  amount  of  potassium 
sulfocyanate  in  the  saliva. 

The  English  observer  Davidson3  in  1841  found  that  patients 
treated  with  mercury  do  not  give  the  thiocyanate  reaction 
in  the  saliva.  This,  of  course,  is  due  to  the  formation  of  a 
colorless  mercury  thiocyanate.  He  also  observed  that  in 
certain  febrile  diseases,  he  obtained  a  negative  test  for  the 
rhodanate.  In  diabetes,  as  well,  the  presence  of  the  sugar 
might  cause,  he  said,  a  diminution  or  a  complete  absence 
of  the  sulfocyanate.  Longet4  found  that  the  amount  of 
KSCN  is  not  dependent  on  the  age,  sex,  diet  or  the  condition 
of  the  nervous  system.  In  salivation,  the  same  author  con- 
tinues, it  would  sometimes  appear  that  potassium  thio- 
cyanate is  absent,  but  this  is  really  not  the  case,  because  the 
saliva   only   needs   concentration   to   get   a   positive   result. 

1  Pavlov:  Die  Arbeit  der  Verdauungsdrusen,  1898. 

2  Eberle:  Physiologie  der  Verdauung,  1838,  p.  32. 

3  Davidson:  London  Med.  Gazette,  1841,  xxix,  p.  338. 
*  Longet:  Loc.  cit. 


Bruylants1  did  not  find  any  thiocyanates  in  the  secretions 
of  gouty  patients  Kriiger1  confirmed  the  findings  of  Longet. 
Grober'  stated  that  those  patients  that  suffer  from  cachexia 
from  any  cause  show  very  little  hydrosulfocyanic  acid  in  the 
saliva. 

The  toxic  effects  of  the  sulfocyanates  have  been  given 
some  attention;  but  the  opinions  advanced  are  very  con- 
flicting. Wohler2  and  Frerichs3  were  of  the  opinion  that  the 
salts  were  not  at  all  toxic  in  quite  large  doses.  Claude 
Bernard,4  Setschenow5  and  Podcopaew8  reported  individually 
that  the  potassium  thiocyanate  has  some  toxic  action. 
Nysten7  quotes  Littre  and  Charles  Robin  as  saying  that  the 
sulfocyanate  of  potassium  is  very  toxic.  Paschkis8  was  also 
of  the  opinion  that  the  toxic  influence  of  this  substance  is 
to  be  reckoned  with.  Guareschi9  found  that  the  rhodanate 
precipitates  many  organic  bases  and  alkaloids,  and  thus 
acts  as  a  kind  of  prophylactic  in  cases  of  poisoning.  When 
Chouppe10  attempted  to  grow  plants  using  saliva  as  a  watering 
agent,  he  found  that  the  growth  rapidly  faded  and  withered. 
Raulin11  noticed  that  in  growing  the  Aspergillus  fumigatus, 
it  was  necessary  to  add  iron  to  the  spraying  water  to  neutralize 
a  poison  in  the  plant,  which  tended  to  prevent  its  growth. 
This  deleterious  substance  was  hydrosulfocyanic  acid. 
The  toxicologist  Witthaus12  in  his  discussion  of  sulfocyanic 
acid  states  that  it  is  poisonous;  very  much  more  so  than  its 
salts. 

1  Loc.  cit. 

I  Wohler:  Gilbert's  Ann.,  1829,  lxii,  p.  271. 

3  Frerichs:  Wagner's  Handworterbuch  der  Phys.,  1964,  iii,  p.  766. 

4  Bernard :  Lecons  sur  les  effets  des  substances  toxiques  et  medica- 
mentenses  Paris,  1857,  p.  386. 

*  Setschenow:  Virchow's  Archiv.,  1857,  xiv,  p.  356. 
'  Podcopaew:  Virchow's  Archiv.,  1878,  xxxiii,  p.  505. 

7  Nysten:  Sulfocyanure,  1878,  p.  15. 

8  Paschkis:  Wiener  mediz.  Jahresber.,  1885,  p.  531. 

•Guareschi:  Introduzione  alio  studio  degli  alcoloidi.    Torino,  1892, 
P-  33- 

10  Chouppe:   Florian,  Gaz.  med.  dc  Paris,  1884,  p.  354. 

II  Raulin:  Gamier  and   Schlagdenhauffen;    Fremy's   Enclyclopedie- 
chimique,  1892,  ix,  p.  195. 

11  Witthaus:  "Organic  and  Inorganic  Chemistry,"  1905,  p.  302. 


30 

Lauder  Brunton1  affirms  that  in  mollusca,  hydrosulfo- 
cyanic  acid  has  the  following  actions:  It  diminishes  reflex 
actions  and  quickens  the  heart  beat;  large  doses  arrest  the 
heart  in  systole. 

Lately2  a  series  of  experiments  was  performed  showing 
the  effects  of  sodium  thiocyanate  on  the  output  of  saliva. 
I  shall  quote  the  following  verbatim : 

''Experiment  X. — A  series  of  experiments  was  then  in- 
stituted to  show  the  effects  of  i  grain  of  sodium  thiocyanate 
taken  internally.  Specimens  were  taken  before,  and  36  and 
80  minutes  after. 

"The  subject  was  conscious  of  a  slightly  stimulating  effect, 
and  showed  an  increased  blood  pressure.  There  was  a 
decrease  in  the  enzyme,  but  the  last  specimen  was  taken  an 
hour  after  a  noon  meal. 

"One  of  the  most  noteworthy  findings  of  this  plot,  which 
has  been  verified  by  a  number  of  other  findings,  is  the  de- 
crease in  the  amount  of  mucin;  but  the  increase  in  the  centri- 
fuged  sediment  was  found  to  be  true  in  the  examination  of 
ten  other  specimens,  as  this  drug  has  a  physiological  effect 
in  reducing  sediment  of  all  kinds  in  the  saliva  as  well  as  the 
urine." 

The  antiseptic  action  of  saliva  in  general,  and  of  the  thio- 
cyanates  specifically,  was  studied  by  many  scientists.  In 
1833,  Kletzinsky3  suggested  that  the  potassium  thiocyanate 
functionated  in  the  saliva  as  an  antiseptic.  Gamier  and 
Schlagdenhauffen4  reported  that  the  thiocyanates  have  some 
bactericidal  properties.  Edinger5  suggested  the  theory  that 
sulfocyanic  acid  builds  various  compounds  with  the  different 
aromatic  radicals  in  the  body,  and  is  thus  enabled  to  protect 
the  body  against  infectious  diseases.  He  found  that  KSCN 
was  antiseptic  for  diphtheria  bacillus,  cholera  bacillus,  and 
the    staphylococcus    pyogenes    aureus.     Another    observer, 

1  Brunton:  Pharmacology,  1878,  p.  114. 

2  Dental  Cosmos,  191 1,  liii,  p.  1297.     Report  of  the  committee. 

3  Heller's  Archiv.,  1833,  P-  39- 

*  Gamier  and  Schlagdenhauffen:  Loc.  cit. 

•Edinger:  Ber.  d.  Freiburger  naturforsch.  Gesel.,  1894,  ix,  p.  27. 


3i 

Martinotti,1  did  not  wish  to  come  to  a  positive  conclusion 
whether  potassium  rhodanate  inhibits  the  development  of 
tuberculosis  in  the  living  organism.  An  Italian  bacteri- 
ologist, Sanarelli2,  found  that  the  saliva  rapidly  destroys 
cultures  of  streptococcus  pyogenes,  staphylococcus  pyogenes 
aureus,  micrococcus  tetragenus,  and  the  cholera  bacillus,  but  it 
has  no  effect  upon  the  diphtheria  bacillus  and  the  pneumococcus. 

Some  dentists,  especially,  have  found  that  the  saliva  due 
to  its  KSCN  content  has  antiseptic  and  bactericidal  prop- 
erties. It  has  been  thought  by  some  of  the  members  of  this 
profession  that  the  potassium  sulfocyanate  prevents  the 
growth  of  plaques  on  the  teeth  and  thus  is  prophylactic  for 
dental  caries.  Michel3  considers  the  rhodanates  as  a  natural 
protective  agent  against  tooth  decay.  Low4  noticed  that 
when  the  thiocyanates  are  present,  the  teeth  are  almost 
always  free  from  caries;  patients  who  have  caries  have  no 
KSCN  or  the  very  faintest  traces  in  their  saliva;  and  he 
further  reports  that,  clinically,  when  he  fed  his  patients  KSCN, 
the  caries  disappeared.  He  expressly  wishes  it  to  be  under- 
stood, however,  that  the  thiocyanate  in  his  opinion  has  no 
antiseptic  action.  Kirk5  has  fully  criticized  Low  in  a  recent 
article  pointing  out  some  of  the  contradictory  phases  of  Low's 
theory.  The  latter,  however,  has  found  support  in  the  work 
of  Waugh6  and  Roberts  who  found  that  potassium  thio- 
cyanate restricts  the  growth  of  the  dental  plaque. 

There  have  been  not  a  few  observers,  on  the  other  hand, 
who  have  found  exactly  opposite  results  to  the  ones  reviewed 
above.  Miller7  stated  that  KSCN  does  not  possess  any 
appreciable  antiseptic  influence  in  the  greatest  strength  in 
which  it  is  found  in  the  human  saliva.  Likewise,  Hugen- 
schmidt8  considered  the  bactericidal  property  of  saliva  very 
slight  indeed,  if  it  is  at  all  present.     Two  French  scientists, 

1  Martinotti:  Centralbl.  f.  Bakt.  u.  Parasitkunde,  1896,  xix,  p.  142. 

2  Sanarelli:  Arch.  ital.  di.  clinic  med.,  1891,  iii,  p.  230. 

3  Michel:  Deut.  Monatsch.  f.  Zahnheilk,  1911,  xxix,  p.  507. 

4  Low:  Loc.  cit. 

6  Kirk:  Dental  Cosmos,  1911,  liii,  p.  1345. 
*  Waugh:  Dental  Cosmos,  1910,  Hi,  p.  420. 

7  Miller:  Dental  Cosmos,  1903,  xlv,  p.  689. 

8  Ilugenschmidt:  Dental  Cosmos,  1896,  xxxviii,  p.  881. 


32 

Nicholas  and  Dubief1  did  not  find  the  thiocyanate  nor  the 
saliva  to  have  any  antiseptic  properties.  Barnes2  obtained 
data  that  contradicted  somewhat  the  results  of  Sanarelli  and 
Edinger.  He  reported  that  the  saliva  has  no  bactericidal  action 
on  the  pneumococcus,  streptococcus  pyogenes,  staphylococcus 
pyogenes  albus,  and  the  influenza  bacillus.  Wounds  in  the 
mouth  heal  rapidly,  he  observed,  due  to  the  leucocytes  that 
are  present  in  the  salivary  secretion. 

Black,3  the  discoverer  of  the  dental  plaque,  denied  ab- 
solutely that  the  saliva  had  any  antiseptic  action.  Oppen- 
heim4  contrary  to  the  findings  of  Smith,4  Auftrecht,4  Michel5 
and  Gruber,4  recorded  that  0.5  per  cent.,  1  per  cent,  and  2 
per  cent,  solution  of  the  thiocyanate  had  no  sterilizing  in- 
fluence upon  bacteria,  especially  of  the  fermenting  kind. 

Seaman  and  Gies6  in  1910  did  not  find  that  biological  pro- 
portions of  the  sulf ocyanates  had  any  retarding  influence  upon 
the  production  of  plaques,  or  upon  the  growth  of  bacteria. 

H.  P.  Pickerill  in  his  Cartwright  Prize  Essay  (191 2)  on  the 
Prevention  of  Dental  Caries  and  Oral  Sepsis,  presents  a  very 
brief  and  very  incomplete  review  of  the  literature  on  the  sulfo- 
cyanates.  He  concludes  the  chapter  rather  vaguely,  and  not 
on  the  basis  of  his  findings,  with  the  following  words:  "On  the 
whole,  we  may  conclude  that  while  undoubtedly  sulfocyanate 
of  potassium  is  a  beneficial  element  in  saliva,  and  one  making 
for  freedom  from  disease,  yet  it  can  not  be  regarded  as  the 
most  important  or  only  factor  in  producing  a  natural  immunity 
to  dentalcaries  or  oral  sepsis."7 

There  are  many  authors  whose  papers  I  have  not  re- 
viewed because  they  simply  stated  what  was  well  known  or 
else  they  merely  sided  with  one  or  another  laboratory  worker 
on  general  principles.  I  shall,  however,  give  a  list  of  them, 
without  any  comment  whatever  at  the  end  of  this  dissertation. 

1  Nicholas  and  Dubief:  Jour,  de  Physiol.,  1878,  i,  p.  979. 

2  Barnes:  Trans.  Chicago  Path.  Soc,  1907-1909,  vii,  p.  249. 

3  Black:  Dental  Digest,  1909,  xv,  p.  603. 

*  Oppenheim:  Hecht  Dental  Cosmos,  1909,  li,  p.  1275. 
6  Michel:  Loc.  cit. 

•  vSeaman  and  Gies:  Dental  Cosmos,  191  o,  lii. 

'  This  dissertation  was  already  in  print  when  Pickerill's  book  became 
available. 


CHAPTER  II. 


THE    FERRIC   CHLORIDE — ETHER   TEST. 

Few  investigators  have  doubted  the  reliability  of  the 
qualitative  test  for  the  thiocyanates  discovered  by  Trcvi- 
ranus.  It  is  known  that  certain  substances  give  a  similar 
coloration  when  treated  with  the  chloride  of  iron;  but  their 
occurrence  in  the  saliva  is  so  rare  that  these  substances  have 
been  excluded  from  any  consideration.  Recently,  Bunting1 
advanced  the  theory  that  the  substance  in  the  saliva  that 
gives  the  red  color  with  ferric  chloride  is  usually  not  a  sulfo- 
cyanate,  but  some  other  substances  whose  presence,  properties 
and  reactions  have  not  yet  been  described.  His  conclusions 
are  based  on  a  series  of  experiments  that  he  has  performed. 
I  shall  give  a  brief  summary  of  his  methods  of  analysis. 

A  few  cubic  centimeters  of  each  saliva  were  placed  in  each 
of  two  25  cc.  Erlenmeyer  flasks;  to  one  of  these  flasks  he  added 
0.1  cc.  of  Njioo  KSCN  solution.  The  salivas  were  then 
evaporated  to  dryness  under  suction  over  a  hot  water  bath. 
Bunting  found  that  upon  treating  these  dried  salivas  with  a 
few  drops  of  ferric  chloride  and  then  shaking  with  ether,  he 
invariably  obtained  a  reddish  coloration  of  the  ether  in  all 
the  control  salivas  (■/.  c,  in  those  to  which  he  had  added  some 
KSCN),  but  he  usually  did  not  get  any  tinting  of  the  ether 
when  he  had  not  put  in  some  thiocyanate. 

I  herewith  attach  a  table  of  his  results  and  his  remarks: 


Water  solutior 

1. 

Ether  solution. 

No. 

Color. 

Per  cent. 

Test. 

Control 

I 

Light  red 

O.OO36 

+ 

+ 

2 

Straw 

O . OO I  6 



+ 

3 

Straw 

O.OOO7 



+ 

4 

Light  red 

O.OO35 



+ 

5 

Dark  lemon 

0.002I 



+ 

6 

Amber  red 

O.OO46 

Faint 

+ 

7 

Dark  red 

O.OO51 

+ 

+ 

8 

Dark  lemon 

O.OO32 

Trace 

+ 

9 

Dark  lemon 

O.OO26 

Trace 

+ 

10 

Lemon 

O.OOO9 

+ 

1 1 

Red 

O.OO43 

Faint 

+ 

12 

Light  red 

OOO34 

Faint 

+ 

1  Bunting:  Dental  Cosmos,  1910,  Hi,  p.  1346. 


34 

"  From  these  results,  a  wide  discrepancy  in  the  two  methods 
is  obvious.  Of  the  five  cases  giving  weak  or  doubtful  tests 
in  water,  but  one  gave  any  test  in  ether.  It  must  be  remem- 
bered that  ferric  sulfocyanate  shows  a  much  more  delicate 
color  test  in  ether  than  in  water,  so  that  KCNS,  if  present  at 
all,  should  be  discernible  in  the  ether  solution.  Of  the  re- 
maining seven  which  gave  a  positive  distinctly  red  color 
reaction,  there  were  but  two  that  gave  a  like  reaction  in  ether. 
These  two  were  distinctly  red  in  their  reaction  and  more 
pronounced  than  in  the  water  solution.  The  remaining  five 
gave  a  negative  or  very  faint  reaction." 
Experimental. 

i.  When  KSCN  solution  is  treated  with  a  few  drops  of 
dilute  hydrochloric  acid  and  a  few  drops  of  3  per  cent,  solu- 
tion of  ferric  chloride,  a  brilliant  red  coloration  is  obtained, 
This  when  shaken  with  ether,  gives  a  cherry-red  tint  to  the 
ethereal  layer  above  the  water.  If  the  thiocyanate  solution 
is  diluted  so  as  to  give  a  light  red  color,  there  will  be  no  red 
coloration  of  the  ether.  Upon  shaking  the  dilute  thio- 
cyanate with  the  ether,  the  following  facts  are  to  be  noted : 

A.  That  the  ether  is  not  tinted.  B.  That  the  color  of  the 
ferric  thiocyanate  in  the  watery  layer  becomes  much  paler. 

The  red  color  in  the  ethereal  solution  of  ferric  thiocyanate 
can  be  readily  caused  to  disappear  by  the  addition  of  a  few 
drops  of  water. 

If  a  few  cubic  centimeters  of  saliva  be  put  into  a  test  tube 
and  treated  with  some  dilute  HC1  and  FeCl3,  a  reddish  colora- 
tion is  invariably  obtained.  Quite  infrequently  one  sees 
the  straw,  lemon  or  dark  lemon  hues  of  which  certain  authors 
speak.  The  reddish  tint  may  be  light  or  quite  dark.  Upon 
shaking  with  ether,  no  matter  what  the  color  may  be,  the 
color  becomes  markedly  paler  without  imparting  any  rosy 
tint  to  the  ethereal  layer. 

2.  Upon  twenty-five  specimens  of  saliva  from  the  mouths 

of  healthy  persons  as  well  as  diseased  (from  the  wards  of  Mt. 

Sinai    Hospital),    I   carried   out   the  following   modification1 

of  the  ferric  chloride  test  for  KCNS  in  saliva  in  ether  solution: 

1  Bunting:  Loc  cit. 


35 


Color  of  ether. 

Pink 
Pink 


Pink 


Five  cc.  of  saliva  were  placed  in  a  round  watch  glass  and 
evaporated  to  dryness  over  a  slowly  steaming  water  bath. 
To  the  dried  residue,  there  were  added  two  drops  of  water 
and  one  or  two  drops  of  ferric  chloride.  This  was  then  well 
stirred  to  make  a  thick  paste.  Five  cc.  of  ether  were  then 
added  and  stirred  well.  The  color  of  the  ether  was  then 
noted. 

Table  I. 

Saliva  No.  Color  with  FeCl3. 

i  Deep  red 

2  Deep  red 

3  Light  red 

4  Light  red 

5  Red 

6  Light  red 

7  Light  red 

8  Light  red 

9  Light  red 
io  Dark  red  Pink 

1 1  Light  red 

1 2  Light  red 

13  Red  Pink 

1 4  Light  red 

15  Deep  red  Pink 

16  Light  red 

1 7  Light  red 

18  Red  Pink 

19  Light  red 

20  Light  red 

2 1  Red  Pink 

22  Light  red 

23  Light  red 

24  Light  red 

25  Light  red 

Of  the  twenty-five  cases  examined  by  the  watch  glass 
method,  eight  gave  a  reddish  tint  to  the  ethereal  layer. 
The  other  seventeen  gave  absolutely  no  color  to  the  ether. 
All  of  them,  upon  being  treated  with  ether  became  lighter 
in  hue,  even  though  in  the  great  majority  of  cases  the  ethereal 
layer  did  not  become  colored. 

3.  Twenty-four  of  these  salivas  were  tested  for  the  thio- 
cyanates   by   the   Erlenmeyer   flask-suction   method,  as  rec- 


36 

ommended  by  Bunting.       The  exact  method  of  procedure 
was  as  follows : 

Into  each  of  two  25  cc.  Erlenmeyer  flasks  were  put  5  cc. 
of  saliva.  To  one  of  these  flasks,  there  was  added  0.1  cc. 
of  a  N/ioo  KSCN  solution.  The  flasks  were  provided  with 
rubber  stoppers  through  which  passed  glass  tubing,  bent  at 
right  angles  outside  the  flasks.  These  glass  tubes  were 
connected  to  a  suction  pump,  and  the  flasks  heated  in  a  water 
bath  whose  temperature  was  kept  at  40  °  C.  The  salivas  were 
evaporated  to  dryness  and  treated  as  follows:  To  each 
flask  were  added  one  drop  of  water  and  one  to  two  drops  of 
FeCl3  solution  and  shaken  with  5  cc.  of  ether.  The  color 
of  the  ether  of  the  two  flasks  containing  the  saliva  was  then 
compared.  The  results  as  they  were  noted  are  given  in 
the  accompanying  table. 

Table  II. 


Saliva  No. 

Color  of  water  solution. 

Ether  so 
Test. 

lution. 
Control. 

I 

Deep  red 

+ 

4- 

2 

Light  red 



— 

3 

Light  red 



— 

4 

Red 

+ 

+ 

5 

Light  red 



— 

6 

Light  red 



— 

7 

Light  red 



4-* 

8 

Light  red 



— 

9 

Deep  red 

+ 

+ 

10 

Light  red 



— 

11 

Light  red 



— 

12 

Red 

+ 

4- 

13 

Light  red 



— 

14 

Deep  red 

+ 

+ 

15 

Light  red 



— 

16 

Light  red 



— 

17 

Red 

+ 

4- 

18 

Light  red 



— 

19 

Light  red 



+  * 

20 

Red 

+ 

+ 

21 

Light  red 



— 

22 

Light  red 



— 

23 

Light  red 



— 

24 

Light  red 



— 

*See  page. 

37 

Of  the  twenty-four  salivas  examined  with  the  suction  and 
control  methods,  seven  gave  positive  results  without  the 
addition  of  any  thiocyanate.  Fifteen  specimens  of  saliva 
gave  not  the  faintest  tint  to  the  ether,  either  in  the  test  flask  or 
in  the  control  flask.  Cases  No.  7  and  19  behaved  somewhat 
out  of  the  ordinary.  These  two  salivas  resemble  the  speci- 
mens examined  by  Bunting. 

Nos.  7  and  19  upon  being  dried  under  suction  at  a  tempera- 
ture of  400  C,  and  then  treated  with  a  drop  of  water  and  two 
drops  of  FeCl3  gave  no  color  to  the  ether  upon  being  shaken 
with  the  latter.  These  same  salivas  gave  a  faint  pink  color 
to  the  ether  in  the  control  flask,  i.  e.y  in  that  flask  to  which 
0.1  cc.  N/100  KSCN  had  been  added. 

Correlation  and  Explanation  oj  the  Foregoing  Findings. 

Undissociated  ferric  thiocyanate  is  soluble  in  ether.  The 
dissociation  of  any  substance  increases  with  the  dilution, 
■j.  c,  the  more  dilute  a  solution,  the  stronger  is  the  dissocia- 
tion and  the  less  the  undissociated  portion  that  is  present. 
At  a  certain  dilution,  a  substance  is  completely  dissociated. 
When  a  dilute  watery  solution  of  ferric  sulfocyanate  (pale 
red)  is  shaken  with  ether,  the  ether  does  not  become  colored 
because  there  is  no  undissociated  thiocyanate  to  enter  the 
ether.  If  we  increase  the  concentration  of  the  thiocyanate 
in  the  water,  the  ether  is  immediately  tinted  rose-red.  There 
is,  therefore,  a  certain  level  or  limit  of  dilution  or  concentra- 
tion of  ferric  thiocyanate  in  water,  below  which  no  un- 
dissociated thiocyanate  remains  and  below  which,  therefore, 
the  ether  will  not  be  colored. 

The  salivas  that  I  have  examined  behaved  quite  similarly 
with  the  dilute  solutions  of  KSCN.  All  the  salivas,  upon 
being  treated  with  dilute  acid  and  a  few  drops  of  ferric 
chloride,  gave  red  colors  varying  in  shade  and  intensity. 
None  of  the  salivas  (without  drying)  imparted  any  tint 
to  the  supernatant  ethereal  layer.  That  is  to  say,  the  amount 
of  KSCN  in  the  saliva  is  so  slight  that,  it  is  completely  dis- 
sociated. 

Eight  of  the  salivas  recorded  in  Table  I  gave,  upon  drying 


38 

and  treatment  with  FeCl3  and  ether,  a  pink  coloration  to  the 
ether.  These  salivas  had  evidently  more  KSCN  than  the 
other  seventeen  eases.  The  solution  formed  by  the  addition 
of  the  few  drops  of  water  and  ferric  chloride  was  not  of 
sufficient  dilution  to  completely  dissociate  the  iron  sulfo- 
cyanate,  and  therefore,  some  of  it  went  into  the  ether  and 
caused  it  to  become  red.  This  explanation  will  hold  for  the 
seven  specimens  of  saliva  that  gave  positive  results  with  the 
suction  method. 

Seventeen  of  the  saliva  specimens  as  recorded  in  Tables 
I  a  ad  II  gave  no  reddish  tint  to  the  ether.  Evidently  the 
thiocyanate  was  present  in  such  very  minute  quantities, 
that  the  addition  of  a  few  drops  of  water  is  quite  capable  of 
completely  dissociating  it  and  thus  preventing  the  ether 
from  assuming  any  rosy  color. 

Specimens  Nos.  7  and  19  are  particularly  interesting. 
These  salivas,  upon  evaporation  on  a  watch  glass  and  testing 
with  ferric  chloride  and  ether,  gave  no  red  color  to  the  ether. 
The  result  was  also  negative  upon  evaporating  under  suction. 
But  when  0.1  cc.  N/100  KSCN  solution  was  added  and  the 
saliva  then  dried  under  lessened  atmospheric  pressure,  a 
positive  result  was  obtained. 

The  explanation  of  these  facts  seems  quite  plain.  The 
amount  of  sulfocyanate  present  in  the  salivary  secretion  was 
just  at  the  limit  of  complete  dissociation.  The  amount  of 
KSCN  that  was  added  was  quite  enough  to  increase  the  con- 
centration of  the  thiocyanates  and  thus  allow  some  of  the 
substance  to  remain  undissociated. 

The  fact,  therefore,  that  frequently  we  do  not  get  a  colora- 
tion of  the  ether  when  shaken  with  dried  saliva  which  had 
been  treated  with  FeCl3  solution  is  no  evidence  of  the  absence 
of  thiocyanates.  In  fact,  according  to  the  light  of  the  disso- 
ciation theory,  it  is  somewhat  a  corroborative  evidence  of 
its  presence.  The  absence  of  the  ether  coloration  is  in  a  way 
a  quantitative  test  of  the  amount  of  the  sulfocyanate  in  the 
saliva.  For  it  is  evident  that  slight  amounts  of  thiocyanates 
will  give  a  negative  result,  while  greater  quantities  will  cause 
a  positive  reaction. 


39 

4.  Substances  that  will  give  a  similar  color  with  ferric  chloride, 
or  that  may  interfere,  in  general,  with  the  sulfocyanatc  test. 

(a)  Neutral  Formates.1, — When  a  neutral  formic  salt  is 
treated  with  dilute  ferric  chloride,  a  deep  red  coloration  is 
produced.  This  color  is  still  retained  when  the  medium  is 
just  acid,  i.  e.,  upon  addition  of  a  few  drops  of  formic  or 
acetic  acid.  One  or  two  drops  of  10  per  cent.  HC1  may  be 
added  without  changing  the  color,  but  a  few  drops  more  of 
this  acid  dispels  the  red  hue. 

I  made  a  solution  of  the  neutral  formate  which  when  treated 
with  dilute  neutral  ferric  chloride  gave  a  red  coloration  of  the 
same  intensity  as  the  average  saliva  when  similarly  treated. 
Of  this  solution  of  the  neutral  formate  (untreated  by  FeCl3), 

1  added  known  amounts  to  saliva.  I  then  treated  the  saliva 
together  with  the  formate  with  FeCl3  solution,  and  noticed 
the    effect    produced.     I    examined    three    salivas.     I     put 

2  cc.  of  each  saliva  into  each  of  six  tubes.  Tube  No.  i,  I 
treated  with  2  drops  of  HC1  and  several  drops  of  FeCl.,. 
Tubes  Nos.  2,  3,  4,  5,  6  were  each  respectively  treated  with 
2,  5,  8,  10  and  15  drops  of  the  neutral  formate  solution,  and 
then  tested  with  the  dilute  ferric  chloride.  The  salivas 
examined  were  of  those  tested  by  Bunting's  suction  process. 

Table  III. 
Salivas  to  which  has  been  Added  Neutral  Formate. 

on  tfi 

a  a 

o  o 

•a  t3 


Bunting 

method. 

a 

CO 

a 

e* 

■0 

0 

a 

01 

73  a 

« 

a 

0 

"o 

(Ng 

<n 

> 

*j 

s 

OJ  •— 
OS    O 

£  - 

~ 

r. 

£ 

8 

0 

£ 

"£~ 

% 

I 

Acid 

+ 

_L 

Light 

red 

+ 

-f 

2 

Acid 





Light 

red 

+ 

4- 

3 

Neutral 

+ 

T 

Light  red 

+ 

+ 

+    Dark  red  + 

+       +      +      Dark  red 
+       +      +      Dark  red 

(6)  Neutral  Acetate. — The  same  experiments  were  per- 
formed with  the  acetates  as  with  the  formate  on  five  samples 
of  saliva.  The  following  table  records  the  results.  A  plu- 
sign  indicates  a  darkening  of  color. 

1  Weston:  Identification  of  Carbon  Compounds,  1907,  p.  15. 

1  The  reaction  in  each  case  was  tested  by  means  of  litmus  paper. 


40 


Table  IV. 

Salivas  to  which  has  been  Added  Neutral  Acetate. 


Bunting 
method. 

a               a 
o               o 

a 

o 

■o»j           -S 

_o 

"o 

(N^2                     lO 

o 

at  u 

s§     i 

Pi 

CD        0 

* 

&       * 

Alkaline 



Light 

red 

+      + 

Alkaline 



Light  Light 

red 

red        + 

Neutral 

+      + 

Light 

Light  Light 

red 

red       red 

Acid 

+      + 

Light 

red 

+          + 

Acid 

—     + 

Light 
red 

+          + 

+  +  + 


+  +  +  + 


+      + 


+  + 


+  + 


+  + 


+    +  +   Dark  red 


+        + 


+  + 


(c)  The  neutral  trichlor acetate  has  the  same  effect  as  the 
neutral  formate  or  neutral  acetate.  Ethyl  acetate  is  not 
colored  red  by  FeCl3. 

(d)  Pyrogallic  acid  gives  a  very  dark  red  color  in  quite 
dilute  solution,  when  treated  with  ferric  chloride.  The 
accompanying  table  will  explain  itself.  The  effect  of  this 
substance  upon  the  thiocyanate  test  in  the  saliva  is  only  of 
theoretical  importance,  since  practically  this  polyphenol 
never  occurs  in  the  oral  secretions.  When  an  excess  of 
potassium  hydroxide  is  added,  the  color  becomes  quite  black. 

Table  V. 
Salivas  to  which  has  been  Added  Pyrogallol. 


Bunting 
method. 


Pi 

Alkaline 
Alkaline 
Neutral 
Neutral 
Neutral 


H     O 


u  o 
-w  O 


—  Light  red 


+    + 


+ 
+ 
+ 
+ 


+  +    +  +  + 


a 
o 

<X> 

+  +  + 
+  +  + 


dark 
dark 


-t-  ■+■     -t-  -r  T  "I"  T  f  UcJXK 

+  +    +  +  +  +  +  +  +  dark 

+  ++  +  +  +  +  +  dark 

+  +        +  +  +  +  +  dark 


(e)   Certain  substances  give  a  violet  or  purplish  coloration 
upon  treatment  with  ferric  chloride.     These  chemical  bodies, 


4' 

should  they  he  present  in  the  saliva  (as  they  sometimes  un- 
doubtedly are),  will  cause  a  darkening  of  the  color  formed 
in  the  sulfocyanate  test,  and  the  amount  of  the  rhodanate 
will  consequently  seem  excessive.  Phenol  and  salicylic  acid 
were  especially  examined.  Either  of  these  substances  upon 
addition  to  saliva,  caused  a  darkening  of  the  hue,  so  that  the 
color  seemed  tawny  red. 

A  dilute  solution  of  the  sodium  salicylate  was  used  in  these 
experiments. 

Table  VI. 
Saliva  to  which  has  been  Added  Salicylate. 


Bunting 

c 

ui 

method. 

J3 

n 

&2 

00 

% 

BO 

a 

0 

a 

a 

o 

a 
o 

u 

C 

o 

H 

-s 

•a 

T3 

T3 

o 

r. 

"o 

h  o 

(N   o 

■* 

>o 

M 

> 

u 

*;      & 

So 

A% 

JS 

J3 

J2 

J3 

a 

■ 

tn           C 

CD  u 

.—  in 

*j 

*j 

♦j 

*J 

<2 

V          0 

£ 

£ 

s: 

^ 

^ 

'^ 

I 

Acid 



Pale  red 

Pale  red 

+ 

_)_ 

+  4- 

+  4- 

2 

Neutral 

4-    + 

Pale  red 

Pale  red 

+ 

4- 

4- 

+  + 

3 

Neutral 



Pale  red 

Pale  red 

Pale  red 

4- 

+ 

4-  + 

4 

Acid 

—  — 

Pale  red 

Pale  red 

+ 

+ 

+  4- 

+  + 

5 

Alkaline 



Pale  red 

Pale  red 

+ 

+ 

+ 

+  4- 

The  addition  of  a  few  drops  of  HC1  caused  a  paling  of  the 
color,  due  to  the  disappearance  of  the  salicylate  shade. 

The  cresols  give  color  reactions  upon  treatment  with  ferric 
chloride.  Orthocresol  gives  a  light  green  color  to  a  dark 
olive  green,  depending  upon  the  dilution.  Para  cresol  gives 
a  pale  blue  color. 

Pyrocatechol  gives  with  FeCl;!  a  dark  green  color  which 
changes  to  violet.  Orsinol  gives  a  violet-blue  color. 
Creosote,  which  has  been  used  in  the  treatment  of  tuber- 
culosis, and  is  used  now  by  dentists  as  a  dental  germicide 
contains  phenol,  cresol,  creasol  and  guaiacol.  With  ferric 
chloride  it  gives  a  brown-red  color. 

With  a  2  per  cent,  phenol  solution,  nearly  similar  results 
were  obtained.  Resorcin  should  also  be  mentioned  here,  as  a 
possible  complicating  substance.  Upon  shaking  the  colored 
phenol  solution  with  ether,  it  becomes  decolorized. 

(/')  Benzoates  and  Succinates. — The  neutral  salts  of  these 
acids    give    a    red    precipitate    with    ferric    chloride.     If    the 


42 

saliva  is  viscid,  mucinous — "ropy" — the  precipitate  will  not 
settle  out  and  will  appear  to  be  diffuse.  These  substances, 
though  presumably  present  in  slight  amounts  in  the  saliva, 
should  be  reckoned  with,  because  many  of  the  vegetable 
sauces  and  canned  goods  are  rich  in  the  benzoates.  The 
color  that  a  very  thick  saliva  assumes  if  a  few  drops  of  the 
neutral  benzoate  is  added,  and  if  it  be  then  treated  with  iron 
chloride,  is  a  dark  brick-red.  The  addition,  however,  of 
several  drops  of  a  10  per  cent.  HC1  tends  to  lighten  the  color. 

(g)  Care  must  be  taken  to  prevent  the  alkaline  carbonates 
or  hydrates  from  interfering  with  the  thiocyanate  test.  With 
ferric  chloride  these  substances  give  an  orange-red  to  a  brick- 
red  precipitate.  Usually  their  presence  is  so  slight  that  no 
fear  need  be  had  of  their  complicating  effect.  The  addition 
of  dilute  HC1  causes  this  precipitate  to  dissolve  and  vanish. 

(h)  Meconic  acid,1  C5H02.(OH).(COOH)2,  is  a  substance 
which  is  peculiar  to  opium  in  which  it  exists  in  combination  with 
a  part,  at  least  of  the  alkaloids.  It  crystallizes  in  small  pris- 
matic needles;  is  acid  and  astringent  to  taste,  loses  its  water 
of  crystallization  at  1200  C,  is  quite  soluble  in  water  and 
alcohol,  sparingly  soluble  in  ether.  With  ferric  chloride, 
it  forms  a  blood-red  color  which  is  not  discharged  by  mercuric 
chloride  or  by  dilute  acids,  but  is  discharged  by  stannous 
chloride  and  by  alkaline  hypochlorides.  This  acid  may, 
theoretically,  be  present  in  salivas  of  patients  suffering  from 
acute  opium  poisoning  or  chronic  morphinomania. 

Table  VII. 
Salivas  to  which  has  been  Added  Meconic  Acid. 


a 

Bunting 
method. 

c 

.0 

3 
O 

in 

a 

U 

09 

S 

■a 

10 

a 
0 

a 
0 

■s 

0 

> 

0 

3 

O 

V)           C 
V            0 

a  0 

~8 

CO 

5 

£ 

1 

Neutral 



Yellow-red  Yellow-red 

Yellow-red 

+ 

+  + 

2 

Neutral 

■ 

Pale  red 

Pale  red 

Pale  red 

+ 

+  + 

3 

Acid 



Pale  red 

Pale  red 

Pale  red 

+ 

+  + 

4 

Alkaline 

+  + 

Dark  red 

Dark  red 

Dark  red 

+ 

+  + 

5 

Acid 



Pale  red 

Pale  red 

Pale  red 

+ 

+  + 

1  Witthaus:  Inorganic  and  Organic  Chemistry,  1905. 


43 

In  the  above  tests  a  very  dilute  solution  of  meconie  acid 
was  used — one  that  with  FeCl3  gave  a  color  reaction  which 
resembled  a  i  :  5000  solution  of  KSCN.  The  quantities 
added  to  each  test  tube  are  indicated  in  the  accompanying 
table.  The  addition  of  a  few  drops  of  the  acid  caused  a 
deepening  of  shade. 

(0  Aceto-acetic  acid  or  diacetic  acid,  CH,.CO.CH2.COOH, 
occurs  in  urine  of  febrile  diseases  and  in  advanced  cases  of 
diabetes.  It  is  conceivable,  a  priori,  that  in  such  conditions 
of  disease,  this  acid  may  be  also  excreted  in  the  saliva.  Bunt- 
ing gave  this  acid  as  an  example  of  a  substance  that  might 
conflict  with  the  thiocyanate  test.  Jones1  without  giving 
any  authority  for  his  opinion,  states  that  diacetic  acid  is 
"ever  present."  This  is  a  somewhat  strange  "finding"  for 
it  is  known  that  this  substance,  when  it  does  occur  in  the 
human  organism,  is  accompanied  by  the  severest  symptoms 
of  metabolic  disturbance.  Together  with  be taoxy butyric 
acid  and  acetone,  it  is  considered  in  the  clinical  pathology 
of  urine,  as  a  sign  of  grave  and  unfavorable  prognostic  im- 
port. One  can  not  imagine  that  this  volatile  substance  should 
calmly  remain  in  the  saliva  after  Bunting's  drastic  treatment. 

Diacetic  acid2  with  ferric  chloride  gives  a  violet-red  or 
Bordeaux-red  color,  which  disappears  upon  addition  of  HC1. 
It  is  soluble  in  ether  to  which  it  gives  a  yellowish  red  color 
not  at  all  similar  to  the  ferric  thiocyanate  in  ether.  The 
color  in  the  ether  disappears  spontaneously  in  twenty-four  to 
forty-eight  hours.  The  fact  that  its  color  persists  upon 
addition  of  hydrochloric  acid,  is  enough  to  differentiate  it 
from  sulfocyanate. 

It  was  thought  advisable  however  to  examine  ten  salivas 
for  diacetic  acid  by  the  Arnold- Lipliawsky  reaction.  The 
reagent  consists  of  two  solutions  (a)  a  1  per  cent,  solution  of 
potassium  nitrite,  (6)  1  gram  of  /?-aminoacetphenon  dis- 
solved in  1 00  cc.  distilled  water  and  about  2  cc.  of  HC1  (con- 
centrated) added  drop  by  drop  until  the  solution,  which  is  at 
first  yellow,  becomes  colorless.  Before  using,  a  and  b  are 
mixed  in  the  ratio  of  1   :  2. 

1  Jones:  Dental  Review,  1911,  xxv,  p.  1167. 
*  Hawk:  Physiological  Chemistry,  1910. 


44 

The  test  is  conducted  as  follows : 

Place  5  cc.  of  filtered  saliva  and  an  equal  volume  of  the 
reagent  in  a  test  tube;  add  a  few  drops  of  concentrated 
ammonia,  and  shake  the  tube  vigorously.  A  brick-red  color 
is  produced.  Take  i  cc.  of  this  colored  solution,  add  10-20 
cc.  HC1,  3  cc.  chloroform,  and  2-4  drops  of  ferric  chloride 
solution,  and  carefully  mix  the  liquids  without  shaking. 
If  the  chloroform  assumes  a  blue  or  violet  color,  the  test  is 
positive  for  diacetic  acid;  if  this  acid  is  absent  the  color  may 
be  yellow  or  light  red. 

In  ten  cases  of  saliva,  tested  by  this  method,  not  a  trace 
of  this  acid  was  found.  In  five  cases  of  diabetic  saliva,  no 
diacetic  acid  was  found  by  this  test.  The  addition  of  a  dilute 
pure  solution  of  this  acid  to  saliva  causes  a  darkening  of 
color. 

Table  VIII. 
Saliva  to  which  has  been  Added  Diacetic  Ester. 


n 

> 

Si 

i 

Bunting 
method. 

"o 

«S                 * 

m         a 
V           0 
h<        0 

V) 

*J  0 
CD  u 

xn 

a 
0  . 

•5-3 

a 
0 

* 

JA 

a 
0 

00 

i 

I 

Acid 

— 

— 

Pale  red 

+ 

+ 

+  + 

+  + 

2 

Neutral 

— 

— 

Pale  red 

+ 

+ 

+  + 

+  + 

3 

Acid 

— 

— 

Yellow-red 

+ 

+ 

+  + 

+  + 

4 

Alkaline 

— 

— 

Yellow-red 

+ 

+ 

+  + 

+  + 

5 

Alkaline 

— 

— 

Pale  red 

T 

+ 

+  + 

+  + 

(j)  Antipyrin  and  Phenacetin. — Two  coal  tar  products  in 
common  use  as  analgesics  and  antipyretics  give  a  red  color 
when  treated  with  the  ferric  chloride  solution.  Experi- 
mentally, upon  addition  of  several  drops  of  solution  of  these 
substances  to  saliva,  and  then  testing  with  ferric  chloride, 
a  deepening  of  shade  was  noticed,  even  with  very  dilute  solu- 
tions.    Salophen  ma)'  here  be  mentioned  as  well. 

(k)  Certain  quinolin  homologues — as  thallin,  ethyl  thallin 
and  kairin  have  been  used  in  medicine  as  antiperiodics  and 
antipyretics.  If  these  substances  are  excreted  unchanged 
in  the  saliva,  they  may  complicate  the  thiocyanate  test,  for 
they  also  give  a  red  coloration  with  iron  chlorid  solution. 


45 

(/)  Sodium  thiosuljatc — in  weak  solution  gives  a  very 
transient  violet-red  color  upon  treatment  with  dilute  ferric 
chloride  solution,  the  thiosulfate  being  oxidized  to  the  sulfate 
state. 

(m)  Glycocoll — aminoacetic  acid — CH2.NH2COOH — is  one 
of  the  amino  acids  produced  by  disintegration  and  decomposi- 
tion of  proteins.  In  dilute  solutions  with  neutral  ferric 
chloride  solution  it  gives  an  intense  red  color.  It  is  not 
soluble  in  ether.  Upon  adding  varying  quantities  of  this 
substance  (in  dilute  solutions)  to  several  samples  of  saliva, 
and  then  testing  with  ferric  chloride,  no  deepening  of  the  red 
shade  was  noticed,  except  in  cases  in  which  the  salivary 
thiocyanate  content  was  very  weak  or  else  when  the  amount 
of  glycocoll  added  was  comparatively  excessive. 

(n)  Amidol. — /»-Diamidophenol  hydrochloride,  is  a 
synthetic  product  used  in  developing  photographic  plates. 
In  extremely  dilute  solutions,  it  gives  an  intense  red  color 
with  ferric  chloride.  The  addition  of  one  or  two  drops  of  a 
very  dilute  solution  of  this  chemical  to  a  few  cubic  centi- 
meters of  saliva,  and  then  treating  with  the  iron  chloride 
induces  an  extreme  red  coloration. 

(o)  Patients  undergoing  a  long  course  of  treatment  with 
mercury  may  not  give  the  thiocyanate  test  in  their  saliva.1 
This  is  no  evidence  that  they  completely  lack  the  rhodanate. 
It  may  be  that  the  mercury  (especially  in  cases  of  salivation) 
so  effects  the  thiocyanate  as  to  produce  the  colorless  mercury 
sulfocyanate. 

5.  Spontaneous  disappearance  0}  the  red  color  in  the  ethereal 
layer. 

(a)  In  all  the  salivas  that  gave  positive  results  by  the 
Bunting's  suction  test  for  thiocyanate  (t.  e.,  those  cases  in 
which  the  ether  was  colored  pink) ,  the  following  phenomenon 
was  observed  The  pink  ethereal  layer,  upon  being  drawn 
off  into  a  clean  test  tube  and  being  allowed  to  stand  from 
5-30  minutes  was  spontaneoulsy,  decolorized  in  each  case. 

(b)  An  ethereal  solution  of  thiocyanate  was  made  by 
shaking    some    absolute    ether    with    a    concentrated    ferric 

1  Davidson:   London  Medical  Gazette,  1841,  xxix,  p.  338. 


46 

thiocyanate  solution.  The  ethereal  layer  was  then  drawn 
off  and  diluted  with  ether,  until  the  pink  color  produced 
resembled  closely  the  color  of  the  ethereal  extracts  obtained 
in  the  positive  cases  of  saliva  by  Bunting's  method.  This 
pale  red  ethereal  solution  was  put  in  several  clean  test  tubes, 
each  well  stoppered,  and  allowed  to  stand  four  to  twenty- 
four  hours.  A  paling  of  color  was  noticed  in  all  of  the  tubes, 
and  some  tubes  were  completely  colorless. 

Upon  several  repetitions,  I  was  able  to  obtain  as  a  starting 
point  a  very  pale  rose-red  ether  which  upon  being  allowed  to 
stand  from  one  to  two  hours  was  entirely  decolorized  spon- 
taneously. 

CONCLUSIONS. 

i.  The  ferric  chloride  colorimetric  test  for  thiocyanates 
in  saliva  is  inexact  and  unreliable. 

2.  A  negative  result  by  the  Bunting  suction  method,  is  no 
evidence  of  the  absence  of  the  sulfocyanate  in  saliva. 

3.  A  positive  result  by  the  Bunting  suction  method  is 
evidence  of  a  comparatively  large  amount  of  sulfocyanate 
in  the  saliva. 

4.  Various  medicinal  substances  and  various  chemical 
compounds  that  are  the  result  of  decomposition  of  proteids 
and  carbohydrates  may,  if  excreted  in  the  saliva,  give  a  very 
marked  red  coloration  when  treated  with  ferric  chloride,  and 
thus  convey  the  impression  that  the  amount  of  thiocyanate 
is  very  large. 

5.  There  is  a  spontaneous  disappearance  of  the  pink  color 
in  the  ethereal  layer  in  the  positive  cases  to  the  Bunting 
suction  method. 


CHAPTER  III. 

COMPARISON    OF    QUANTITATIVE    METHODS. 

I  shall  eschew  an  extensive  discussion  of  the  colorimetric 
methods  for  the  quantitative  determination  of  the  thio- 
cyanates. A  resume  of  such  proceedings  as  have  been  in 
vogue  during  the  last  century  was  given  in  the  review  of  the 
literature  on  thiocyanates.  I  am  convinced  that  to  rely 
upon  colorimetric  analyses  will  lead  one  to  very  inaccurate 
results.  There  is  too  much  of  the  personal  element  in  the 
comparison  of  the  intensity  of  two  shades  of  color. 

There  are  two  procedures  for  the  rhodanate  determination, 
that,  though  tedious  and  requiring  precise  accuracy  in 
manipulations,  will  give  exact  figures.  These  methods  are 
(i)  gravimetric,  (2)  iodometric. 

Jacubowitsch1  whose  extensive  studies  on  saliva  were  con- 
sidered classical  was  one  of  the  first  to  suggest  a  gravimetric 
method  for  the  determination  of  thiocyanates.  He  was 
especially  interested  in  the  salivary  secretions,  but  his  method 
can  be  used,  with  slight  modification,  in  examining  most  of 
the  tissues  and  fluids  of  the  body.  He  collected  750  cc. 
saliva,  and  extracted  several  times  with  alcohol.  The 
extract  was  filtered  and  the  filtrate  evaporated.  The  nearly 
dry  residue  was  distilled  with  phosphoric  acid.  In  this  way 
he  oxidized  all  the  sulfur  to  the  sulphate  ion,  and  he  then 
precipitated  this  with  barium  hydrate  and  barium  nitrate. 
The  sulfate  of  barium  was  collected  on  an  ashless  filter  paper, 
filtered,  dried,  ignited  and  weighed. 

Munk,2  modified  this  method  by  first  precipitating  the 
thiocyanate  with  silver  nitrate  and  then  oxidizing  it  by 
fusion  with  soda  and  the  nitrate  of  sodium.  This  method 
will  be  described  in  detail  a  little  later  (vide  -infra). 

The  gravimetric  method  suggested  by  Bruylants3  seems 
to  me  to  be  very  unwieldy  and  not  as  accurate  as  the  other 

1  Jacubowitsch:  "De  Saliva,"  Inaugural  Dissert.,  Dorpat,  1850. 
J  Munk:  Deut.  Med.  Woch.,  1869,  lxix,  p.  427;  1877,  lxxvi,  p.  350; 
also  Arch.  f.  d.  ges.  Physiol.,  1895,  lxi,  p.  620. 

3  Bruylants:  Maly's  Jahres.  d.  Tierchem.,  1888,  xviii,  p.  134. 


48 

methods.     I  shall  simply  mention  it  here;  it  has  been  described 
with  some  detail  in  the  first  chapter. 

In  1894,  Lang1  suggested  a  titration  method  for  the  de- 
termination of  the  thiocyanates.  Two  processes  of  titration 
are  requisite  by  this  method.  The  first  one  determines  the 
amount  of  chloride  plus  thiocyanates  according  to  Volhard;2 
the  second  determines  the  amount  of  chlorides  alone,  ac- 
cording to  Mohr.3  The  difference  between  these  quantities 
gives  the  thiocyanate  figure. 

The  iodometric  method  was  first  suggested  by  Rupp  and 
Schied4  and  was  improved  by  Thiel5  and  Rupp.6  In  1906 
Edinger  and  Clemens7  applied  this  method  in  the  analyses 
of  biological  fluids  and  tissues.  The  method,  as  I  followed  it, 
is  detailed  by  Paul  Meyer8  in  his  extensive  treatise  on  the 
analysis  of  urine. 

Of  all  these  methods,  those  of  Munk  and  Rupp  and  Schied 
were  chosen  as  giving  the  most  accurate  results.     A  com- 
parison of  their  degree  of  accuracy  was  then  attempted. 
Description  of  Methods. 

1.  Munk's  Gravimetric  Method.9 — The  fluid  to  be  examined 
is  filtered.  If  the  substance  is  solid,  it  is  extracted  with  water 
for  twenty-four  hours,  and  the  extract  then  filtered.  The 
solid  portion  is  washed  several  times  on  the  filter  paper  with 
warm  water.  The  filtrate  is  treated  with  some  soda  and  then 
evaporated  on  the  water  bath.  This  is  then  extracted  several 
times  with  alcohol,  and  filtered.  The  filtered  alcoholic 
extract  is  evaporated,  and  the  residue  dissolved  in  water  and 
again  filtered.  The  filtrate  is  acidified  with  dilute  nitric 
acid  and  the  thiocyanate  precipitated  with  silver  nitrate. 
The  solid  silver  chloride  and  silver  thiocyanate  are  collected 

1  Lang:  Arch.  f.  exp.  path.  u.  pharm.,  1894,  xxxiv,  p.  253. 

2  Volhard:  Liebig's  Ann.,  1877,  cxc,  p.  1. 

3  Paul  Meyer:  In  C.  Neuberg's  "Der  Harn,"  1911,  i,  p.  651. 

4  Rupp  and  Schied:  Ber.  deut.  chem.  Ges.,  1902,  xxxv,  p.  219. 
8  Thiel:  Ber.  deut.  chem.  Ges.,  1902,  xxxv,  p.  2766. 

8  Rupp:  Chem.  Zentral.,  1905,  ii,  p.  1288. 

7  Edinger  and  Clemens:  Zeitsch.  f.  klin.  Med.,  1906,  lix,  p.  223. 

8  Meyer:  In  C.  Neuberg's  "Der  Harn,"  1911,  i,  p.  651. 
8  Borcher:  Rcpcrt.  d.  anal,  chem.,  1881,  iv,  p.  130. 


49 

on  an  ashless  filter  paper,  washed  carefully  and  dried  at 
iOO°  C.  The  precipitate  together  with  the  filter  paper  are 
fused  in  a  silver  crucible  with  soda  and  sodium  nitrate. 
The  excess  of  nitric  acid  is  gotten  rid  of  by  adding  hydro- 
chloric acid  solution  and  evaporating.  The  remainder  is 
dissolved  in  water,  filtered  and  the  filtrate  precipitated  with 
barium  chloride.  After  allowing  the  precipitate  to  sediment 
for  twenty-four  hours,  it  is  collected  on  an  ashless  filter 
paper,  dried,  ignited  and  weighed.  From  the  amount  of 
barium  sulphate,  thus  obtained,  one  can  easily  calculate  the 
quantity  of  thiocyanate  present. 

2.  Rupp,  Schied  and  Thicl  lodomelric  Method. — The 
principle  of  this  method  consists  in  the  fact  that  sulfocyanate 
solutions  treated  with  bicarbonate,  decolorize  large  amounts 
of  iodine,  cyanogen  iodide  being  formed, 

CNSK  +  4l2  +  4H20   =  H2S04  +  6HI  +  KI  +  CNI. 

The  process  is  finished  in  four  hours  at  ordinary  tempera- 
ture. Upon  acidifying  with  hydrochloric  acid  solution 
carefully,  the  potassium  iodide  is  changed  to  potassium 
chloride  and  hydriodic  acid  and  this  acts  on  the  cyanogen 
iodide  to  form  hydrocyanic  acid.  The  whole  process  can  be 
expressed  in  the  following  equation: 

CNSK  +  3I2  +  4H20  =  H2S04  +  5HI  +  KI  +  CNH, 
i.  e.,  one  molecule  of  thiocyanate  is  equivalent  to  six  mole- 
cules of  iodine.     The  analysis  is  conducted  as  follows: 

Reagents  used: 

1.  Nitric  acid,  1  per  cent. 

2.  Silver  nitrate,  3  per  cent. 

3.  Infusorial  earth,  clean,  washed  in  acid. 

4.  Sodium  bicarbonate,  C.  P. 

5.  Potassium  iodide,  C.  P. 

6.  iVi/10  iodine  solution. 

7.  Hydrochloric  acid,  10  per  cent. 

8.  N  1/10  sodium  thiosulphate  solution. 

9.  Starch  solution,  2  per  cent. 

The  liquid  to  be  examined  is  filtered.  If  it  is  solid,  it  is 
macerated  and  extracted  with  water  for  twenty-four  hours. 


5Q 

The  extract  is  then  filtered.  In  order  to  remove  any  albu- 
mins that  may  be  present,  the  extract  is  heated  and  the 
precipitated  proteins  filtered  off.  The  clear  filtrate  is 
acidified  with  nitric  acid  and  an  excess  of  silver  nitrate  is 
added.  In  order  to  cause  complete  sedimentation,  some 
infusorial  earth  is  added.  It  is  now  filtered  under  suction 
on  a  filter  paper  stuck  in  a  perforated  platinum  cone.  Care 
must  be  observed  that  the  filtrate  is  clear.  The  collected 
precipitate  is  transferred  to  a  wide-necked  glass  container 
by  means  of  water.  3  grams  of  the  bicarbonate  of  soda  are 
added  to  alkaline  reaction.  Then  3  grams  of  solid  potas- 
sium iodide  are  added  and  the  solution  is  shaken  slightly 
until  it  is  clear.  N  1/10  iodine  solution  is  then  added  until 
a  permanent  brown  color  is  formed.  This  is  then  shaken 
slightly  and  allowed  to  stand  in  a  dark  place  for  four  hours. 
After  acidifying  very  carefully,  with  10  per  cent,  hydrochloric 
acid  solution  a  few  cubic  centimeters  of  the  starch  solution 
are  added,  which  is  freshly  prepared.  This  is  finally  titrated 
with  N  1  j  10  sodium  thiosulfate  solution  until  the  blue  color 
just  disappears. 

The  two  methods  were  tested  in  duplicate  on  pure  solutions 
of  potassium  sulfocyanide  and  on  biological  tissues  and  fluids 
to  which  had  been  added  known  amounts  of  the  thiocyanate. 
The  following  is  a  list  of  the  substances  upon  which  analyses 
were  conducted: 

1.  A  pure  solution  of  potassium  thiocyanate. 

2.  Urine  to  which  was  added  a  known  amount  of  the  thio- 
cyanate. 

3.  Fifty  grams  of  meat  to  which  was  added  a  known  amount 
of  thiocyanate. 

4.  Saliva  (250  cc.)  to  which  was  added  a  known  amount  of 
thiocyanate. 

The  data  found  are  set  forth  in  the  accompanying  table 
(Table  I).  In  my  hands  the  iodometric  method  gave  better 
results.  It  is  not  as  long  nor  as  complicated  as  the  gravi- 
metric method;  the  sources  of  error  and  the  amount  of  error 
are  less;  it  takes  less  time  to  carry  it  out;  four  or  five  analyses 
can  be  easily  run  simultaneously. 


5i 


Table 

I. 

Comparison  of  Results 

by  Iodometric  and  Gravimetric  Analyses. 

Substance  examined  to      Wt. 

Iodometric 

Gravimetric. 

which  had  been  added  a      in 

Amount 

Amount 

known  amount  of  KSCN.  gins. 

found. 

Error. 

found. 

Error. 

Distilled  water       500 

O . O408 

0.0002 

O . O4065 

O.OOO35 

Meat                          50 

O.O3814 

O.OO296 

OO3765 

O   OO345 

Meat                          50 

OO375 

O.OO36 

OO3823 

O.OO287 

Meat                          50 

OO395 

O.OO16 

O.O3844 

O.OO266 

Meat                          50 

O.O3782 

O.OO328 

OO3727 

O.OO383 

Urine                        500 

O . O45 I 

O.OO4* 

O • O463 

O.OO52* 

Urine                       500 

OO432 

0.002I* 

O.O458 

O.OO47* 

Saliva                      250 

O.O447 

O.OO36* 

OO432 

0.002I* 

I  analyzed  the  following  biological  tissues  and  fluids  for 
thiocyanates  to  determine  whether  the  rhodanates  are 
normally  present  and  if  so,  to  what  extent: 

1.  A  liter  of  human  urine. 

2.  500  grams  of  bovine  liver. 


500  cc.  of  human  saliva. 
500  grams  of  beef  meat. 

Table  II. 
KSCN  in  Various  Biologic  Fluids  and  Tissues. 


Weight. 

Tissues. 

Grams. 

Amount  of  KSCN.f 

Urine 

IOOO 

O.O262 

Bovine  liver 

500 

O.OO47 

Human  saliva 

500 

O.OI28 

Beef  meat 

500 

O.O 

I  have  found,  as  many  other  investigators  have  previously 
done,  that  the  urine  and  saliva  contain  varying  amounts  of 
the  rhodanate.  The  bovine  liver,  according  to  my  analysis 
contains  4.7  mg.  per  500  grams  of  the  tissue.  Beef  meat  does 
not  contain  any  thiocyanate. 

♦Increase. 

fin  all  the  analyses  recorded  in  this  dissertation,  the  quantity  of  thio- 
cyanate* was  always  calculated  and  recorded  as  potassium  thiocyanate. 


CHAPTER  IV. 

THE    DISTRIBUTION    OF    THE    THIOCYANATES    IN    THE    ANIMAL 

BODY. 

The  salts  of  sulfocyanic  acid  have  been  found  in  fluids 
of  the  animal  body,  other  than  in  the  saliva.  Musso1  found 
the  thiocyanates  in  milk,  Leared2  in  the  blood,  Gscheidlen3 
in  the  urine,  Nencki4  in  the  gastric  juice,  and  Muck5  in  the 
nasal  and  conjunctival  secretions.  De  Souza6  stated  that 
it  is  present  in  the  blood,  pancreatic  juice  and  bile.  Fen- 
wick7  also  implied  that  the  bile  contains  some  rhodanate. 
Grober8  and  Longet9  on  the  other  hand,  denied  that  the 
thiocyanates  are  present  in  fluids  other  than  saliva.  Bruy- 
lants10  found  the  salts  of  sulfocyanic  acid  in  milk,  ox  gall, 
in  the  fluid  of  a  hydrocele  and  in  cystic  fluid  of  the  abdomen. 

No  investigator  has  ever  systematically  determined  the 
distribution  of  thiocyanic  salts  in  the  various  organs  and 
tissues  of  the  body.  Vague  statements  are  present  in  the 
literature  on  this  subject,  as  to  the  rhodanate  content  of 
various  species  of  animals,  but  I  have  met  no  exact  figures 
relating  to  the  amount  of  this  substance  in  the  different 
animal  tissues.  It  was  deemed  advisable,  therefore,  before 
proceeding  any  further,  to  determine  the  amount  of  sulfo- 
cyanates  present  in  the  various  organs  of  a  dog. 

General  Description  of  Experiments. — The  organs  and 
tissues  of  six  dogs  were  analyzed.  The  first  four  dogs  used 
had  been  experimented  on  by  other  workers  in  the  labora- 

1  Musso:  Berichte  f.  physiol.  Chem.,  1877,  vii,  p.  168. 

2  Leared:  Proc.  Roy.  Soc.  London,  1869,  xviii,  p.  16. 

3  Gscheidlen:  Arch.  f.  d.  ges.  Physiol.,  1877,  xiv,  p.  401. 

4  Nencki:  Berichte  d.  d.  chem.  Ges.,  1895,  xxviii,  p.  1318. 

5  Muck:  Munch,  med.  Woch.,  1900,  xlvii,  p.  1168. 

•  De  Souza:  Journal  of  Physiol.,  1907,  xxxv,  p.  332. 
7  Fenwick:  Brit.  Med.  Jour.,  1882,  i,  p.  397. 

•  Grober:  Deut.  Arch.  f.  klin.  Med.,  1901,  lxix,  p.  243. 

•  Longet:  Traitc  de  Physiologie,  1868,  i,  p.  191. 

10Bruylants:  Bull,  de  l'Acad.  de  Medicin  Belgique,  1888,  xxi,  p.  147. 


53 

tory.  The  experiments,  however,  that  were  performed  on 
these  animals  could  have  produced  no  radical  changes  in  the 
animals.  The  first  two  dogs  had  several  ounces  of  blood 
taken  away  from  them,  and  the  second  pair  of  animals  had 
been  transfused  with  blood  taken  from  other  dogs.  The 
fifth  and  sixth  dogs  were  perfectly  normal  animals.  Special 
care  was  taken,  as  will  be  seen  later  on,  to  determine  the 
health  condition  of  these  last  two  dogs. 

The  dogs  were  bled  to  death  from  the  right  femoral  arteries. 
Cocaine  was  injected  over  the  site  of  this  blood  vessel.  A 
general  anaesthetic  was  never  used.  The  animal  showed  no 
signs  of  suffering.  All  the  blood  was  collected  in  a  wide 
mouthed,  glass  container,  and  it  was  defibrinated  as  soon  as 
collected.  Particular  care  was  taken  to  exsanguinate  the 
animals  completely.  The  pancreas,  spleen,  kidneys,  liver, 
gall-bladder  and  bile,  heart,  brain,  salivary  glands,  testicles 
and  muscle  of  thigh  were  taken  out  in  the  order  named,  and 
were  put  separately  in  stout,  glass-stoppered,  wide-mouthed 
glass  bottles.  Care  was  observed  that  the  bile  did  not  ooze 
out  and  vitiate  the  results.  Rubber  gloves  were  worn  and 
were  rinsed  in  a  running  stream  of  distilled  water  after  the 
removal  of  each  organ.  Analysis  of  all  the  tissues  was 
begun  immediately  after  their  removal  from  the  body. 

Experiment  i. — A  female  dog,  weighing  15.5  kilos,  was  used 
in  this  experiment.  It  was  noticed  that  the  dog  was  nervous 
and  irritable.  It  was  bled  to  death  on  January  11,  1912, 
from  the  right  femoral  artery.  The  blood  clotted  very 
slowly.  The  exsanguination  was  complete.  No  points  of 
bleeding  were  noticed  upon  opening  the  abdomen.  The 
various  tissues  enumerated  above  were  removed.  The 
heart,  when  opened,  contained  in  its  right  ventricular  com- 
partment, two  whip  shaped  worms  four  or  five  inches  long. 
They  were  carefully  preserved  and  sent  to  the  pathologist 
who  pronounced  the  parasites  to  be  the  filaria  hematis. 

The  amount  of  potassium  sulfoeyanide  found  in  the  various 
tissues  are  charted  in  Table  I. 


54 
Table  I. 


Weight. 

Amt.  of  KSCN 

Tissue. 

Grams. 

Mg. 

Bile 

25 

None 

Blood 

955 

52.86 

Brain 

79 

None 

Heart 

92 

None 

Kidneys 

87 

None 

Liver 

393 

29.84 

Muscle 

200 

None 

Pancreas 

30 

None 

Salivary  glands 

11 

None 

Spleen 

32 

None 

Experiment  2. — A  young  male  pup  (9-10  months  old), 
weighing  7.48  kilos,  was  bled  to  death  on  Jan.  31,  1912. 
The  dog  had  not  finished  his  first  dentition  and  his  testicles 
were  as  yet  undescended.  The  bleeding  was  quite  complete. 
The  accompanying  table  contains  the  analytic  data  (Table  II) . 


Table  II. 

Weight. 

Amt.  of  KSCN 

Tissue. 

Grams. 

Mg. 

Bile 

12 

None 

Blood 

598 

27.32 

Brain 

72 

None 

Heart 

53 

None 

Kidneys 

5i 

None 

Liver 

332 

17.82 

Muscle 

200 

None 

Pancreas 

21 

None 

Spleen 

22 

None 

Experiment  3. — A  male  dog  weighing  9.34  kilos  was  bled  to 
death  on  February  5,  191 2.  The  procedure  was  identical 
with  the  ones  described  above.  The  animal  had  been  trans- 
fused several  times  with  the  blood  of  another  dog.  The 
same  precautions  were  observed  and  the  same  organs  removed 
and  immediately  analyzed.  It  was  thought  advisable  to  tie 
off  the  small  and  large  intestines,  to  remove  them  separately 
and  to  analyze  these  organs  and  their  contents  for  sulfo- 
cyanate.  The  small  intestine  was  tied  off  at  the  duodenum 
and  ileo-caecal  junction.  Of  the  large  intestine,  the  portion 
between  the  caecum  and  rectum  was  used. 


55 


The  analytical  data  are  given  in  Table  III. 
Table  III. 

Tissue. 

Bile 

Blood 

Brain 

Heart 

Small  intestine 

Large  intestine 

Kidneys 

Liver 

Muscle 

Pancreas  and  spleen 

Experiment  4. — The  various  organs  and  tissues,  enumerated 
in  the  preceding  experiments  (together  with  the  small  and 
large  intestines  and  stomach)  were  removed  from  a  male  dog 
weighing  11.3  kilos  on  February  13,  1912.  The  dog  was 
thoroughly  exsanguinated  from  the  left  femoral  artery  under 
local  cocaine  anaesthesia.  The  analytical  findings  are  as 
follows : 

Table  IV. 


Weight. 

Ann.  of  KSCN, 

Grams. 

Mg. 

37 

4-3 

752 

32.56 

63 

None 

69 

None 

635 

7-4 

235 

5  8 

53 

None 

217 

19.  2 

200 

None 

60 

None 

Weight. 

Amt.  of  KSCN. 

Tissue. 

Grams. 

Mg. 

Bile 

28 

7  65 

Blood 

65O 

4234 

Brain 

87 

None 

Heart 

65 

None 

Intestine  (small) 

67O 

22 .  72 

Intestine  (large) 

285 

14  97 

Kidneys 

66 

None 

Liver 

278 

39  5 

Muscle 

200 

None 

Pancreas  and  spleen 

79 

None 

Stomach  and  contents 

210 

None 

Experiment  5. — In  this  experiment  and  the  following,  an 
attempt  was.  made  to  determine  the  normality  of  the  dogs 
before  their  organs  were  analyzed.  The  dogs  selected  were 
to  all  appearance  full  grown,  vigorous  and  healthy.  They 
were  kept  in  cages  devised  by  Prof.  Gies.1  The  urine  daily 
voided  was  collected  and  analyzed  for  sulfocyanates.  The 
same  was  done  with  the  feces. 

1  Gies:  Amer.  Jour.  Physiol.,  1905,  xv,  p.  403. 


56 

The  dogs  were  fed  daily  at  10  a.m.  the  following  mixture: 
Meat1  15  grams,  cracker  meal  4  grams,  lard  3  grams,  bone-ash 
1  gram,  and  water  35  cc.  per  each  kilo  of  weight  of  the  dog. 

The  dogs  were  kept  in  the  cages  for  six  days,  until  we  were 
satisfied  that  they  were  healthy,  normal  dogs.  On  the 
seventh  day  the  animals  were  bled  to  death. 

The  first  dog  was  a  female  weighing  12. 1  kilos.  It  was 
bled  to  death  on  February  27,  1912,  from  the  right  femoral 
artery.     The  analytical  results  are  the  folio  wing :  (Table  V). 

Table  V. 


Weight. 

Amt.  of  KSCN. 

Tissue. 

Grams. 

Mg. 

Bile 

9 

3-0 

Blood 

765 

17.8 

Brain 

58 

None 

Heart 

65 

None 

Intestine  (small) 

589 

96 

Intestine  (large) 

175 

5-5 

Kidneys 

73 

None 

Liver 

280 

10. 7 

Muscle 

200 

None 

Pancreas  and  spleen 

64 

None 

Stomach  and  contents 

128 

None 

Experiment  6. — The  preceding  experiment  was  duplicated 
on  a  male  dog.  weighing  11.75  kilos.  It  was  also  bled  to 
death  on  February  27,  1912,  after  a  feeding  period  of  seven 
days.     The  analytical  figures  are  as  follows :    (Table  VI). 


Table  VI 

Weight. 

Amt.  of  KSCN, 

Tissue. 

Grams. 

Mg. 

Bile 

4 

None 

Blood 

635 

20.35 

Brain 

61 

None 

Heart 

67 

None 

Intestine  (small) 

595 

9-4 

Intestine  (large) 

H7 

8.7 

Kidneys 

52 

None 

Liver 

230 

12.2 

Muscle 

200 

None 

Pancreas  and  spleen 

58 

None 

Stomach  contents 

172 

None 

s:  Amer.  Jour.  Physiol., 

1901,  V, 

P-  235- 

57 

Study  of  the  Sulfocyanate  Content  of  the  Salivary  and  Olln  t 
Glands  oj  the  Body. 

According  to  some  authorities,  all  the  rhodanates  in  the 
animal  organism  are  produced  in  the  salivary  glands. 
Gscheidlen1  caused  a  drainage  of  the  saliva  away  from  the 
mouth  and  found  that,  while  the  secreted  saliva  still  contained 
sulfocyanate,  the  blood  and  the  urine  ceased  to  show  the 
presence  of  this  substance.  Brubaker2  localized  the  forma- 
tion of  the  thiocyanate  in  the  parotid  gland;  he  states  that 
the  submaxillary  and  sublingual  bodies  have  nothing  to  do 
with  the  formation  of  this  substance. 

I  analyzed  the  salivary  glands  of  the  ox  and  the  dog. 
In  250  grams  of  ox  gland  tissue,  I  found  only  a  slight  amount 
of  sulfocyanate,  viz.,  10.23  mg-  It  *s  quite  impossible 
to  state  the  amount  of  thiocyanate  in  the  salivary  bodies 
of  the  dog.  The  total  weight  of  the  glands  in  a  dog  is  very 
small,  so  that  I  was  unable  to  obtain  a  figure  for  one  dog. 
The  expedient  was  suggested  of  collecting  the  glands  of  five 
or  six  dogs  and  analyzing  en  masse. 

Various  other  glands — duct  and  ductless — of  the  animal 
organism  were  analyzed.  The  ox  salivary  glands,  the  ox 
thyroid,  the  ox  liver,  the  calf  thymus,  the  dog's  salivary 
glands,  the  dog's  spleen,  the  dog's  pancreas,  the  dog's  liver 
and  the  dog's  testicles  were  each  quantitatively  analyzed 
for  potassium  sulfocyanide.  In  the  case  of  the  salivary 
glands  and  testicles  of  dogs,  an  analysis  was  run  on  the  col- 
lected glands  from  six  dogs. 

The  data  of  these  analyses  are  given  in  Table  VII. 


Table  VII. 

Weight. 

Amt.  of  KSCN 

Tissue. 

Animal. 

Grams. 

Mg. 

Liver 

Dog 

393 

28.94 

Liver 

Ox 

500 

4-7 

Pancreas 

Dog 

30 

None 

Salivary  glands 

Dog 

65 

None 

Salivary  glands 

Ox 

250 

10.23 

Spleen 

Dog 

32 

None 

1  Gscheidlen:  Loe.  eit. 

2  Brubaker:  Text  Book  of  Physiology,  1908,  p.  156. 


58 
Table  VII  (Continued) . 


Weight. 

Amt.  of  KSCN. 

Tissue. 

Animal . 

Grams. 

Mg. 

Thymus 

Calf 

370 

None 

Thyroids 

Ox 

605 

None 

Testicles 

Dog 

IO7 

None 

Blood 

Ox 

5OO 

7.6 

Bile 

Ox 

500 

9-7 

Feces 

Man 

235 

12.8 

Table  VIII  shows  the  location  of  the  thiocyanates  in  the  vari- 
ous tissues  and  fluids  of  the  animal  body. 

Table  VIII. 


Salivary 
Saliva      Glands 


Man  + 
Dog  — 
Ox      .  .  .  . 


+ 


Blood 

+ 
+ 


Bile 

+ 
+ 


+ 
+ 


Intestine  Urine     Feces 
+  + 

+  +  + 


CONCLUSIONS. 

i.  The  salts  of  sulfocyanic  acid  are  found  in  certain  fluids 
and  tissues  of  the  body  outside  of  the  saliva  and  salivary 
glands. 

2.  The  liver  seems  to  be  the  gland  in  the  body  which  con- 
tains most  of  the  thiocyanate. 

3.  The  thiocyanate  of  the  liver  is  excreted  through  the  bile 
into  the  small  and  large  intestines,  where  quite  perceptible 
quantities  were  found.  The  stomach  contents  showed  no 
trace  of  the  sulfocyanate  in  dogs  normally  fed.  When  how- 
ever, sodium  sulfide  is  given  the  stomach  contents  showed 
traces  of  sulfocyanate. 

4.  The  blood  contains  appreciable  quantities  of  the 
rhodanate. 

5.  Of  the  glands  of  the  body,  the  liver  and  the  salivary 
bodies  (ox)  show  the  presence  of  the  thiocyanate.  I  was 
unsuccessful  in  my  attempt  to  demonstrate  the  presence  of 
sulfocyanate  in  dog's  salivary  glands.  The  spleen,  the 
pancreas,  the  thymus,  the  thyroid  and  the  testicles  do  not 
contain  any  thiocyanate. 


CHAPTER  V. 

THE     METABOLISM     AND    EXCRETION     OF    SULFOCYANATES. 

Having  determined  the  distribution  of  the  thioeyanates 
in  the  animal  organism,  a  study  was  made  of  the  excretion 
and  metabolism  of.  this  substance. 

It  was  surmised  by  Grober1  that  potassium  sulfocyanate 
is  produced  in  the  organism  by  decomposition  of  proteins. 
De  Souza2  found  that  after  feeding  acetonitrile  to  dogs,  the 
sulfocyanic  salts  were  present  in  large  amounts  in  the  urine, 
saliva  and  serum.  Kabdebo3  concluded  from  his  investiga- 
tions that  acetonitrile  lessens  the  oxidation  of  sulfur,  and 
that  the  neutral  sulfur  unites  with  the  acetonitrile  to  form 
sulfocyanic  acid.  Willianen4  demonstrated  to  his  own 
satisfaction  that  upon  feeding  glycocoll  and  other  amino 
acids  to  rabbits,  he  caused  an  excretion  of  sulfocyanic  acid  in 
the  animals'  urine,  which  otherwise  failed  to  show  the  presence 
of  sulfocyanate.  Diena5  recently  studied  the  effects  of  the 
ingestion  of  rhodalzid  (an  albumen-sulfocyanate  compound 
prepared  by  Nerking)  upon  the  excretion  of  sulfocyanates 
in  the  saliva,  gastric  juice,  pancreatic  juice  and  bile.  He 
made  fistulae  to  these  organs  in  animals  and  collected  the 
juices  after  giving  tablets  of  rhodalzid.  He  found  that  the 
saliva  showed  very  marked  traces  after  the  administration 
of  the  substance.  The  pancreas  and  stomach  showed  smaller 
traces.  He  reported  the  findings  of  Kondo  (Japan)  that  the 
rhodalzid  causes  increase  in  purin  bases  output. 

Having  these  results  in  mind,  it  was  proposed  to  study 
first  the  normal  excretion  of  thioeyanates  in  the  dog. 

Dogs  V  and  VI  were  fed  as  has  been  previously  described, 
their  urine  and  feces  were  daily  collected,  and  the  amount  of 
thiocyanate  determined. 

I  found  that  the  urine  contained  small  amounts  of  sulfo- 
cyanate, while  the  feces  were  quite  rich  in  this  substance. 

1  Grober:  Deut.  Arch.  f.  klin.  Med.,  iooi,  lxix,  p.  243. 

2  De  Souza:  Journal  of  Physiology,  1901,  xxxv,  p.  332. 

3  Kabdebo:  Maly's  Jahr.  d.  Tierchemie,  1907,  xxxvii,  p.  403. 

*  Willianen:  Bioehem.  Centralbl.,  1906,  v,  p.  477. 

*  Diena:  Bioehem.  Zeit.,  1912,  xxxix,  p.  13. 


6o 


The  animals  were  fed  on  meat,  cracker  meal,  lard  and  bone 
ash.  Several  samples  of  a  mixture  of  these  substances  were 
analyzed  for  sulf ocyanates ;  none  was  found. 

The  accompanying  tables  record  the  quantities  of  thio- 
cyanates  found  respectively  in  the  urine  and  feces. 


Date. 

Feb.    2 1 

22 


23 

24 

25 
26 

27 


Date. 

Feb.    21 


23 
24 

25 
26 

27 


Table  i.— Dog  V. 


Urine. 
Cc. 

49O 

450 

460 

3IO 

275] 
500  \ 

350  J 


Amount  of 
KSCN  in  urine. 

Mg. 

8-3 

7-4 
11. 4 


Table  2.— Dog  VI. 

Amount  of 

KSCN  in  urine. 

Mg. 

7-5 


63 


Amount  of 

KSCN  in  feces. 

Mg. 

15-7 
15-2 

29.8 


Amount  of 

KSCN  in  feces. 

Mg. 

12-5 


12  .6 


17.6 


27-5 


Effect  of  the  Administration  of  Powdered  Sulfur  upon  the 
Excretion  of  Sulf  ocyanates. 
Two  male  dogs,  full  grown  and  normal  to  all  appearances 
were  used  in  these  experiments.  The  dogs  were  put  in  their 
cages  on  February  28,  19 12  and  were  fed  daily  at  9.30  a.m. 
Their  meal  consisted  of  meat  15  grams,  cracker  meal  4  grams, 
lard  3  grams,  bone  ash  1  gram  and  water  35  cc.  per  each 
kilo  of  weight  of  the  animals.  Their  excreta  were  collected 
daily,  weighed  and  measured,  and  then  analyzed  for  the 
thiocyanate.  On  March  4,  191 2,  after  having  observed  the 
dogs  for  five  days  and  found  them  normal,  the  feeding  of 
sulfur  was  begun.     The  first  day  each  of  the  dogs  was  given 


6i 


2  grams  of  flowers  of  sulfur,  administered  in  a  ball  of  meat. 
Great  care  was  taken  that  none  of  the  sulfur  should  be  lost. 
If  the  animal  refused  to  take  the  meat-ball  from  the  hand, 
its  mouth  was  opened  and  the  meat  with  the  sulfur  was 
forcibly  pushed  down  the  pharynx  of  the  animal. 

On  March  5th,  3  grams  were  given  to  each  animal;  on 
March  6th,  4  grams;  on  March  7th,  5  grams;  March  8th,  6 
grams;  March  9th,  7  grams;  March  10th,  8  grams. 

The  feces  and  urine  were  daily  analyzed  for  thiocyanates. 
The  results  are  to  be  found  in  Tables  3  and  4. 


Table 

3- 

—Dog 

VII.- 

-Weight 

11.36  Kilos. 

Amount  of 

Amount  of 

Amount  of 

S 

given. 

Urine. 

KSCN  in  urine.     Feces. 

KSCN  in  feces. 

Date. 

Grams. 

Cc. 

Mg. 

Grams 

Mg. 

Feb. 

29 

550 

3    4 

42 

78 

Mar. 

1 

450 

3 

7 

45 

8 

5 

2 

3IO 

3 

2 

49 

8 

9 

3 

560 

4 

1 

40 

8 

•7 

4 

540 

4 

3 

36 

9 

.  1 

5 

2 

280 

4 

3 

47 

8 

•7 

6 

3 

47O 

3 

9 

85 

9 

5 

7 

4 

42O 

4 

2 

105 

12 

3 

8 

5 

370 

3 

2 

62 

10 

.  2 

9 

6 

350 

4 

4 

58 

10 

4 

10 

7 

42O 

3 

7 

55 

10 

.0 

1 1 

8 

380 

4 

3 

59 

11 

•5 

Table 

4 

—Dog 

VIII. 

— Weight 

8.65  Kilos. 

Amount  of 

Amount  of 

Amount  of 

S  given. 

Urine. 

KSCI 

Feces. 

KSCN  in  feces. 

Date. 

Gram: 

Cc. 

Mg. 

Grams. 

Mg. 

Feb.    29 

220? 

37 

8.4 

Mar.      i 

34oi 

5-7 

44 

7  9 

2 
3 

320/ 

2  70  s 

5-9 

39 
4i 

10 
9 

2 

7 

4 

300 

32 

42 

9 

5 

5 

2 

170 

3-7 

38 

10. 

2 

6 

3 

320 

3  5 

47 

9 

6 

7 

4 

250 

3  9 

65 

9 

7 

8 

5 

340 

3  6 

57 

9 

3 

9 

6 

37o 

3  6 

55 

8. 

8 

10 

7 

330 

3  8 

49 

10. 

5 

1 

1 

8 

320 

3  5 

47 

9 

4 

On  March  11,  191 2,  dogs  VII  and  VIII  were  bled  to  death 


62 

from  their  left  femoral  arteries  under  local  cocaine  anesthesia. 
The  organs  and  tissues  were  removed  and  analyzed  for  sulfo- 
cyanate.  The  analytical  data  for  the  organs  of  the  two  dogs, 
are  given  in  Tables  5  and  6. 

Table  5.— Dog  VII. 


Weight. 

Amount  of  KSCN. 

Tissue. 

Grams. 

Mg. 

Bile 

17 

3-2 

Blood 

653 

27.6 

Brain 

63 

None 

Heart 

51 

None 

Intestine  (small) 

253 

10.3 

Intestine 

(large) 

127 

8.7 

Kidneys 

63 

None 

Liver 

258 

25.8 

Muscle 

200 

None 

Spleen 

22 

None 

Pancreas 

12 

None 

Stomach  contents 

I IO 

None 

Table  6.- 

-Dog  VIII. 

Weight. 

Amount  of  KSCN. 

Tissue. 

Grams. 

Mg. 

Bile 

22 

4.O 

Blood 

580 

22.5 

Brain 

60 

None 

Heart 

73 

None 

Intestine  (small) 

295 

n. 2 

Intestine 

(large) 

55 

5-4 

Kidneys 

62 

None 

Liver 

297 

21 .6 

Muscle 

200 

None 

Pancreas 

22 

None 

Spleen 

27 

None 

Stomach  contents 

128 

None 

It  will  be  seen  from  the  Tables  3-6  that  the  feeding  of 
elementary  sulfur  has  no  effect  upon  the  excretion  of  the 
sulfocyanate  substances.  The  average  daily  output  of  sulfo- 
cyanic  salts  in  the  urine  and  feces  was  the  same  before  and 
after  the  administration  of  flowers  of  sulfur.  Neither  did 
these  experiments  show  that  sulfur  produces  an  increase  in 
the  (so  to  speak)  stored  up  thiocyanates  in  the  various  organs 
and  tissues  that  contain  this  substance. 


63 


Effect  of  Administration  of  Sodium  Sulfide  upon  the  Elimina- 
tion of  Sulfocyanatcs, 

Two  dogs  whose  excreta  were  analyzed  daily  for  five  and 
eleven  days  respectively  were  fed  according  to  their  weight 
with  meat,  lard,  cracker  meal,  bone  ash  and  water. 

Before  beginning  to  administer  the  sulfide  to  these  animals, 
an  attempt  was  made  to  determine  which  dosage  will  produce 
the  least  amount  of  discomfort  to  the  animals,  and  especially 
which  dosage  will  not  cause  vomiting;  for,  when  the  sodium 
sulfide  enters  the  stomach,  it  is  acted  upon  by  the  hydro- 
chloric acid  of  the  gastric  juice  and  sulfuretted  hydrogen  is 
produced  which  will  cause  emesis. 

A  third  dog  was  taken  and  0.5  gram  sodium  sulfide  was 
given  him  in  a  gelatin  capsule.  He  promptly  vomited. 
Next  day  before  feeding  this  dog  a  capsule  containing  0.2 
gram  sodium  sulfide  was  given.  The  dog  licked  up  his  food. 
After  half  an  hour  there  was  noticed  marked  eructation  of 
hydrogen  sulfide.  The  vomiting  was  very  slight.  It  was 
decided,  therefore,  to  use  0.15  gram  of  sodium  sulfide  for  the 
dogs  IX  and  X. 

On  March  20,  191 2,  dogs  IX  and  X  were  each  given  0.15 
gram  sodium  sulfide  in  gelatin  capsules.  They  ate  their  food 
heartily  and  did  not  seem  to  suffer  any  discomfort.  They 
were  fed  daily  in  like  manner,  0.15  gram  sodium  sulfide  for 
five  days.  The  urine  and  feces  were  collected  daily  and 
analyzed  for  sulfocyanate.  The  daily  urine  and  feces 
analyses  are  given  in  Table  7  and  8. 

Table  7. — Dog  IX. — Weight  6.94  Kilos. 


Aim 
Date. 

Na2S  given. 
Gram. 

Urine. 
Cc. 

KSCN  in  urine. 
Mg. 

Feces. 
Grams. 

KSCN  in  feces 
Mg. 

iar.    16 

"       17 

"       18 

320l 
340 
2IO  J 

8-3 

27  I 

25  t 

9-6 

19 
20 

220  \ 
250  j 

5  5 

24  1 
27   \ 
30  J 

14  7 

21 

O     I5 

I90 

2.9 

35 

7.2 

"       22 

O.I5 

3IO 

46 

32 

7-5 

23 

O.I5 

370 

4  4 

30 

7.8 

24 

O.I5 

300 

3  9 

33 

8.3 

"      25 

O.I5 

220 

4.2 

29 

8-5 

64 


Table  8. — Dog  X. — Weight  8.2  Kilos. 


Amt. 

Na2S  given. 

Urine. 

KSCN  in  urine. 

Feces. 

KSCN  in  feces 

Data. 

Gram. 

Cc. 

Mg. 

Grams. 

Mg. 

Mar.    10 

250  ' 

25] 

II 

27O 

7-4 

22    > 

18.7 

'         12 

29O  J 

27  J 

'         13 

30O 

32  1 

'         14 

280 

8.2 

39 

16.3 

'         15 

330. 

45  J 

'         16 

'         17 
'         18 

340 
370 
330 

I2.3 

32? 
4i5 
37  X 
32  i 

IO.5 
12.2 

19 

220 

20 

2IO 

39 

6.1 

21 

O. 

15 

36O 

4-3 

43 

6.7 

22 

O. 

15 

340 

4-7 

37 

7-5 

23 

O. 

15 

3IO 

4-5 

35 

7-4 

'         24 

O. 

15 

29O 

4-5 

28 

7.8 

'         25 

O. 

15 

280 

4-7 

32 

7.6 

On  March  25,  1912,  the  two  dogs  were  bled  to  death  and 
their  tissues  and  organs  analyzed  for  thiocyanate.  The 
usual  precautions  described  previously  were  taken.  It 
was  found  in  both  cases  necessary  to  bleed  the  animals  from 
both  femoral  arteries,  because  the  blood  clotted  very  easily 
before  complete  exsanguination.  The  blood  was  quite  dark. 
The  results  of  the  analyses  are  reported  in  Tables  9  and  10. 


Table  9. — Dog  IX. 


Tissue. 

Weight. 
Grams. 

Amount  of  ] 
Mg. 

Bile 

Blood 

Brain 

29 

508 

70 

3-7 

38.8 

None 

Heart 

62 

None 

Small  intestine 

328 

20.6 

Large  intestine 
Kidneys 
Liver 
Muscle 

152 

65 
236 
200 

16.2 
None 
27.0 
None 

Pancreas 

24 

None 

Spleen 

28 

None 

vStomach  contents 

131 

3-5 

65 
Tahle  io. — Dog  X. 


Weight. 

Amount  of  KSCN 

Tissue. 

Grams. 

Mg. 

Bile 

25 

None 

Blood 

467 

4238 

Brain 

56 

None 

Heart 

63 

None 

Small  intestine 

289 

17-5 

Large  intestine 

172 

10.2 

Kidneys 

75 

None 

Liver 

267 

22.  7 

Muscle 

200 

None 

Pancreas 

30 

None 

Spleen 

28 

None 

Stomach  contents 

126 

4,3 

The  feeding  of  sodium  sulfide  to  dogs  produces  no  very 
marked  effects  upon  the  thiocyanate  content  or  excretion. 
It  was  noticed,  however  (see  Tables  7  and  8),  that  there  was  a 
slight  rise  in  the  sulfocyanate  excretion  in  the  feces,  espe- 
cially in  the  fourth  and  fifth  day  of  the  feeding.  The  stomach 
contents  in  both  dogs  showed  traces  of  the  thiocyanate. 
This  may  have  been  caused  by  a  regurgitation  of  duodenal 
contents  into  the  stomach.  One  should  be  very  sceptical 
in  deducing  any  conclusion  from  this  finding  purporting  to 
show  that  sodium  sulfide  is  changed  in  the  stomach  to  thio- 
cyanate. On  the  contrary  the  stomach  as  well  as  the  intestine 
should  be  considered  the  excretory  mechanism  for  the  thio- 
cyanate salts. 

Effect  of  the  Administration  of  Taurine  upon  the  Elimination 
and  Distribution  of  Thiocyanatcs. 
On  March  28,  191 2,  dogs  XI  and  XII  weighing  respectively 
8.4  kilos  and  8.55  kilos  were  put  in  separate  cages  and  were 
fed  daily  as  described  previously  for  a  period  of  seven  days. 
On  April  4,  191 2  each  dog  was  given  0.1  gram  taurine  in  a 
gelatine  capsule.  On  succeeding  consecutive  days,  the 
dogs  were  given  0.2  gram,  0.4  gram,  1.0  gram,  1.0  gram. 
The  dogs  were  bled  to  death  from  the  right  femoral  arteries 
on  April  9,  191 2.  The  usual  precautions  were  adopted  and 
the  various  tissues  and  organs  removed  and  analyzed. 


66 


The     analytic  data  for   the   daily    urine    and    feces    are 
given  in  Tables  n  and  12. 


Apr. 


Date. 

Mar.  29 
30 
3i 
1 
2 
3 
4 
5 
6 

7 
8 


Table  ii. — Dog  XI. — Weight  8.4  Kilos 

Amt.  of  taurine.    Urine.     KSCN  in  urine.    Fee 
Grams.  Cc.  Mg.  Gra 


O.  I 

0.2 
O.4 
O.8 
I  .O 


11. 4 


12. 1 


8-3 


6.2 

5-5 


KSCN  in  feces. 
Mg. 


19-5 


22.5 

16.4 

8-5 
8.2 

9    1 

9  7 


Table  12. — Dog  XII. — Weight  8.55  Kilos. 


Date. 

Mar 


Amt.  of  taurine.  Urine. 


Grams. 


Apr. 


29 
30 

31 
I 
2 

3 
4 
5 
6 

7 


o.  1 

O.  2 
O.4 
0.8 
I    O 


KSCN  in  urine. 
Mg. 


7-5 


I2.0 


149 

4-5 
4-7 
5-3 
5  5 


KSCN  in  feces. 
Mg. 


16.9 


17.4 


14.2 

9-4 

9  7 

9.0 

10.4 


Table  13. — Dog  XI. 


Weight. 

Amount  KSCN. 

Tissue. 

Grams. 

Mg. 

Bile 

18 

None 

Blood 

475 

28.5 

Brain 

64 

None 

Heart 

73 

None 

Small  intestine 

305 

15-4 

67 
Table  13. — Dog  XI  {Continued). 


Tissue. 

Weight. 
Grams. 

Amount  KSCV 
Mg. 

Large  intestine 

l62 

9  7 

Kidneys 

74 

None 

Liver 

275 

'5    1 

Muscle 

200 

None 

Pancreas 

27 

None 

Spleen 

30 

None 

Stomach  contents 

207 

None 

Table  14.- 

-Dog  XII. 

Weight. 

Amount  KSCN. 

Tissue. 

Grams. 

Mg. 

Bile 

28 

None 

Blood 

505 

24.6 

Brain 

70 

None 

Heart 

68 

None 

Small  intestine 

327 

17.  2 

Large  intestine 

155 

7-5 

Kidneys 

69 

None 

Liver 

244 

12.7 

Muscle 

200 

None 

Pancreas 

25 

None 

Spleen 

28 

None 

Stomach  contents 

195 

None 

Taurine,  an  amino  sulfonic  acid  produces  no  noticeable 
effect  upon  the  excretion  or  distribution  of  sulfocyanate  in 
the  animal  organism.  In  this  respect  it  behaves  altogether 
unlike  glycocoll  (according  to  Willianen).  The  bile  in  both 
dogs  failed  to  reveal  the  presence  of  sulfocyanate;  the  in- 
testines and  liver  contained  relatively  the  usual  amount  of 
the  rhodanate. 


Effect  of  the  Administration  of  Thiourea  upon  the  Excretion  of 
Sulfocyanates. 
Two  dogs  were  kept  in  the  cages  several  days  to  determine 
their  normal  sulfocyanate  output.  Thiourea  was  then 
administered  to  them  in  meat  balls.  The  doses  given  were 
5.0  grams,  5  grams,  7.5  grams,  10  grams  and  15  grams  on 
consecutive  days.  The  dogs  were  bled  to  death  on  April 
15,  19 1 2.     The  results  will  be  found  in  Tables  15-17 


68 


Table 

Date. 

Apr.    10 
ii 

12 

13 
14 
15 


15 

Thiourea. 
Grams. 


5-0 

7-5 

10. o 


Dog  XIII.— Weight  7.92  Kilos. 


Urine. 
Cc 

290) 

340$ 

285} 

240$ 

2Io), 

200^ 


KSCN  iu  urine. 
Mg. 

5-3 


4-7 


Feces.    KSCN  in  feces. 
Mg. 


l6.2 


4-9 


19.4 


On  April  15th  the  dog  was  given  15  grams  of  thiourea. 
Table  16. — Dog  XIV. — Weight  8.04  Kilos. 


Thiourea. 

Urine. 

KSCN  in  urine. 

Feces. 

KSCN  in  fe 

Date. 

Grams. 

Cc. 

Mg. 

Grams. 

Mg. 

Apr.    10 

270' 

36] 

II 

5-0 

360 

\           8.7 

32 

17.4 

12 

5-0 

380. 

28  J 

13 

5-0 

27O  1 

37  I 

"          14 

7-5 

l6o 

7-5 

54 

20.2 

"          15 

10.0 

I70. 

52  J 

On  April  15th  dog  was  given  15  grams  thiourea. 
The  data  for  the  analysis  of  the  tissues  of  dogs  XIII  and 
XIV  will  be  found  in  Table  17. 


Table  17. 

Dog  XIII. 

Dog. 

XIV. 

Weight. 

Amt.  KSCN. 

Weight. 

Amt.  KSCN 

Tissue. 

Grams. 

Mg. 

Grams. 

Mg. 

Bile 

30 

4.6 

20 

3-7 

Blood 

470 

20.  7 

465 

I9.4 

Brain 

69 

None 

72 

None 

Heart 

67 

None 

65 

None 

Small  intestine 

285 

20.3 

305 

17.6 

Large  intestine 

l6o 

7-1 

I46 

8-3 

Kidneys 

72 

None 

63 

None 

Liver 

263 

17-5 

185 

22.8 

Muscle 

200 

None 

200 

None 

Pancreas 

21 

None 

17 

None 

Spleen 

27 

None 

20 

None 

Stomach  contents 

I70 

None 

222 

None 

Thiourea  under  the  conditions  of  these  experiments  does  not 
produce  any  effect  on  the  amount  of  thiocyanate  eliminated 
from  or  distributed  through  the  body. 


69 

Effect  of  the  Administration    oj    Acetonitrile    upon    the  Excre- 
tion of  Sulfocyanatcs. 

The  feeding  to  a  dog,  weighing  10.77  kilos,  of  0.5  gram  of 
acetonitrile  daily,  caused  a  marked  exacerbation  in  the  sulfo- 
cyanate  output  in  the  urine.  The  blood  also  upon  analysis, 
gave  a  very  high  thiocyanate  figure.  The  results  of  the 
acetonitrile  experiment  will  be  found  in  Tables  iS  and  19. 

Table  18. — Dog  XV. — Weight  10.77  Kilos. 


Amt.  KSCN 

Amt.  KSC 

Acetonitrile. 

Urine. 

in  urine. 

Feces 

in  feces. 

Date. 

Grams. 

cc. 

Mg.        Grams 

Mg. 

Apr.    15 

2  20 

2.8 

52 

IO.7 

16 

24O 

3-4 

28 

8.0 

"    17 

0    5 

230 

3-3 

17 

8.2 

18 

0    5 

270 

4-4 

48 

9     1 

"    19 

0-5 

300 

63 

42 

8.9 

"     20 

OS 

380 

6.7 

43 

8-5 

"          21 

0    5 

24O 

8.9 

27 

8.4 

Table  19. 

—Dog  XV. 

Weight. 

Amt.  KSCN. 

Tissue. 

Grams. 

Mg. 

Bile 

16 

6-5 

Blood 

560 

47.O 

Brain 

72 

None 

Heart 

75 

None 

Small  intestine 

340 

21  5 

Large  intestine 

144 

10.6 

Kidneys 

64 

None 

Liver 

287 

18.4 

Stomach  contents 

142 

None 

Effect  of  the  Administration  of  Thioacetic  Acid  upon  the  Distri- 
bution of  Sulfocyanate  in  the  Animal  Body. 
Dog  XVI,  male,  weighing  13. 45  kilos,  was  used  in  this  ex- 
periment. Thioacetic  acid  is  very  evil  smelling  and  very 
poisonous.  The  dog  was  given  daily  6  to  8  drops  of  thioacetic 
acid  in  a  gelatin  capsule.  The  dog  vomited,  but  he  always 
licked  up  the  vomitus  so  that  none  of  the  acid  was  lost.  He 
was  bled  to  death  on  the  fifth  day.  The  analysis  of  the  organs 
did  not  show  any  variation  from  the  usual  amount  of  sulfocy- 
anates. 


70 


Effect  of  Administration  of  Alanin  upon  the  Excretion  of  Sulfo- 

cyanates. 

An  attempt  was  made  to  corroborate  the  findings  of  Willianen 
who  stated  that  amino  acids,  like  glycocoll,  caused  an  increase 
in  the  sulfocyanate  output  in  the  urine.  The  effect  of  the  ad- 
ministration of  alpha- amino-propionic  acid  will  first  be  discussed. 

A  dog,  weighing  7.85  kilos,  was  fed  from  2  to  5  grams  alanin 
from  April  24  to  29.  It  was  found  that  there  was  a  marked 
increase  in  the  sulfocyanate  elimination  in  the  \  rine.  The 
amount  of  thiocyanate  in  the  feces  remained  constant.  Table 
20  shows  the  daily  KSCN  output  in  the  urine  and  feces. 

Tabus  20.— Dog  XVII.— Weight  7.85  Kilos. 


Date. 

Alanin. 

Grams. 

Urine. 
Cc. 

Amt.  KSCN 

in  urine. 

Mg. 

Feces. 
Grams. 

Amt.  KSCN  in 
feces. 
Mg. 

Apr.    19 

220  } 

28] 

20 

225   f 

7-7 

29   J 

12.8 

'         21 

2IO  J 

42  J 

'         22 

270] 

37] 

'         23 

.... 

230  \ 

8-3 

43 

13  -7 

'         24 

2 

270  J 

35  J 

'         25 

3 

250 

2-5 

30 

5-5 

'         26 

4 

245 

5-7 

32 

5-3 

'         27 

5 

360 

5-2 

4i 

6.2 

'         28 

5 

270 

8.4 

40 

5-o 

'         29 

5 

340 

8.8 

57 

5-4 

On  April  29,  1912,  the  dog  was  bled  to  death.  The  analyses 
of  the  tissues  and  organs  (see  Table  21)  did  not  reveal  any  in- 
crease in  the  sulfocyanates  in  the  tissues. 

Table  21. —Dog  XVII. 


Weight. 

Amt.  KSCN 

Tissue. 

Grams. 

Mg. 

Bile 

21 

6.2 

Blood 

4IO 

21.8 

Brain 

79 

None 

Heart 

58 

None 

Small  intestine 

348 

25-4 

Large  intestine 

61 

10.3 

Kidneys 

45 

None 

Liver 

243 

18.7 

Muscle 

200 

None 

Pancreas  and  Spleen 

48 

None 

Stomach  contents 

105 

None 

7i 

Effect  of  the  Administration  of  Glycocoll  upon  th*    Excretion  of 
Sulfocyanates. 

Willianen  used  5-10  grams  of  amino  acetic  acid  in  his  ex- 
periments, and  he  reported  a  marked  increase  in  the  sulfocy- 
anate  elimination.  I  used  only  0.5  to  1.5  grams,  and  have 
found  that  with  this  dosage  no  effect  was  produced  on  the 
thiocyanate  excretion  or  distribution. 

The  results  found  are  reported  in  Tables  22  and  23. 

Table  22 — Dog  XVIII. — Weight  9.24  Kilos. 


Glycocoll. 

Urine. 

Amt.  KSCN 
in  urine. 

Amt.  KSCN 
Feces.           in  feces. 

Date.               Grams. 

Cc. 

Mg. 

Grams.             Mg. 

Apr.    16         0.50 

2IO 

30 

34] 

II 

17            O.75 

200 

3  4 

22    [           20.5 

a 

l8             I    OO 

420 

2.9 

55  J 

H 

19             1.25 

340 

4-5 

11}  ■'« 

II 

20             I . 50 

370 

3-5 

Table 

23  — 

Dog  XVIII. 

Weight 

Amt.  KSCN 

Tissue. 

Grams. 

Mg. 

Bile 

None 

Blood 

5IO 

29  3 

Brain 

56 

None 

Heart 

71 

None 

Kidneys 

75 

None 

Small  intestine 

385 

27-5 

Large  intestine 

163 

17.2 

Liver 

302 

No  Analysis 

Spleen 

19 

None 

Pancreas 

24 

None 

Muscle 

200 

None 

Stomach  contents 

170 

None 

The  Effect  of  Preventing  the  Saliva  from  entering  the  Stomach, 
upon  the  sulfocyanate  output. 

A  male  bull  dog  weighing  10.4  kilos  was  kept  in  a  cage  for 
several  days  to  determine  his  state  of  health.  He  was  found 
to  be  entirely  normal. 

On  April  20,  191 2,  the  dog  was  put  under  ether  anaesthesia, 
and,  using  all  aseptic  and  antiseptic  precautions,  an  incis  on 


72 

was  made*  in  the  left  side  of  the  neck,  the  esophagus  was  ex- 
posed and  was  cut  transversely.  The  upper  and  lawer  portions 
were  sutured  to  the  skin. 

The  wound  did  not  suppurate.  The  siliva  was  collected  on 
cotton  swathings  tied  around  the  dog's  neck;  no  saliva  could 
possibly  enter  the  stomach.  The  dog  was  fed  twice  daily 
through  a  stomach  tube  inserted  through  the  opening  in  the 
neck ;  the  food  consisted  of  milk,  cracker  meal,  bone  ash,  beef 
extract,  Witte's  peptone  and  gelatin. 

The  dog  was  daily  put  in  a  holder  and  the  urine  was  collected 
when  voided.  In  this  way  825  cc.  of  urine  were  collected  in  a 
period  of  eight  days.  The  analysis  showed  that  even  with  the 
absence  of  saliva  from  the  stomach,  the  urine  still  showed  the 
normal  traces  of  sulfocyanate.  In  370  cc.  urine  there  were  4.2 
mgs.  KSCN;  in  230  cc.  urine,  2.7  mgs.  KSCN:  in  225  cc.  urine 
2.3  mgs.  KSCN. 

The  dog  died  on  May  2,  1912.  The  blood,  bile  and  liver 
were  analyzed.  In  340  grams  of  blood  I  found  15.5  mgs. 
KSCN;  in  272  gms.  of  liver  there  were  17.8 mg.  KSCN,  and  in 
27  grams  bile  there  were  5.4  mg.  KSCN.  The  saliva  which 
was  collected  through  the  opening  in  the  gullet,  did  not  con- 
tain any  rhodanate. 

Effect  of  Fasting  upon  the  output  of  Sulfocyanates. 

A  male  dog,  weighing  8.46  kilos  was  deprived  of  food  for  a 
period  of  seven  days.  He  was  given  water  ad  libidum.  His 
urine  was  daily  collected  and  analyzed  for  sulfocyanates.  The 
daily  amount  of  rhodanate  eliminated  did  not  vary  from  the 
average  for  the  other  dogs.  On  the  seventh  day  of  fasting, 
the  dog  was  exsanguinated  from  the  left  femoral  arteries.  The 
analysis  of  the  tissues  shows  that  fasting  diminishes  the 
amount  of  sulfocyanates  in  the  body.  The  Intestinal  tract  did 
not  contain  any  rhodanate.     (See  Table  24.) 

*  I  wish  to  express  my  indebtedness  to  Dr.  Morris  Hirsch  Kahn  of 
Mount  Sinai  Hospital  for  the  performance  of  this  operation  and  for  his 
cooperation  in  sending  me  numerous  specimens  of  saliva  from  his  ward 
patients. 


73 


Table  24.— Doc.  XX. 

Weight. 
Tissue.                                       (".111. 

Amt.  KSCN 

Bile                                          19 
Blood                                      455 
Brain                                        55 

4-7 

153 

None 

Heart                                      60 

None 

Small  intestine                      180 

None 

Large  intestine                       50 
Kidneys                                  45 
Liver                                      180 

None 
None 

12.4 

Pancreas  and  Spleen             32 
Stomach  contents                  85 

None 
None 

CONCLUSIONS. 

1.  The  sulfocyanates  are  normally  present  in  various  parts 
of  the  animal  body. 

2.  The  sulfocyanates  are  excreted  in  the  urine  and  feces 

3  The  production  and  elimination  of  sulfocyanates  in  the 
urine  are  not  dependent  upon  the  amount  of  thiocyanate  pres- 
ent in  the  saliva.  Dog  saliva  apparently  is  free  from  sulfo- 
cyanate  while  the  urine  invariably  contains  it. 

4.  The  ingestion  of  amino  acids  (like  alanin),  and  of 
nitriles  (like  acetonitrile)  increases  the  excretion  and  distribu- 
tion of  thiocyanates  in  the  body. 

5.  Thiocyanate  is  apparently  produced  from  protein  in  the 
body.  The  results  for  the  fasting  dog  harmonize  with  this 
conclusion. 

6.  The  ingestion  of  sulfur,  sodium  sulfide,  thioacetic  acid, 
thiourea  and  taurine  did  not  increase  the  output  of  sulfo- 
cyanates. 


CHAPTER  VI. 

THE    TOXIC    EFFECTS    OF    POTASSIUM    SULFOCYANATE- 

Lauder  Brunton1  found  that  solutions  of  potassium  sulfo- 
cyanate,  when  administered  to  mollusca,  diminish  the  reflex 
actions,  but  have  little  effect  on  the  excitability  of  the  nerves. 
A  small  dose  of  this  salt  somewhat  quickens  the  cardiac 
action,  a  large  dose  stops  the  heart  in  diastole,  and  if  it  is 
directly  applied  to  the  heart,  the  stoppage  is  permanent. 

Kletzinski2  suggested  quite  some  time  ago  that  the  sulfo- 
cyanate  of  potassium  is  toxic  to  bacterial  life,  and  its  function 
in  the  saliva  is  bactericidal.  Nysten  quotes  Litre  and  C. 
Robin  that  potassium  thiocyanate  is  very  toxic.  Wurtz, 
on  the  other  hand,  mentions  Woehler  and  Frerichs  as  his 
authorities  for  the  statement  that  potassium  sulfocyanate 
is  not  at  all  poisonous.  Claude  Bernard,  Setschenow  and 
Podcopaew  were  of  the  opinion  that  this  thiocyanate  is  very 
slightly  toxic;  Paschkis,  however,  thinks  that  its  toxicity 
is  quite  noticeable. 

Nerking3  has  synthesized  an  albumen  rhodanate  which  he 
called  rhodalzid.  Lehman4  and  Diena5  studied  this  com- 
pound and  found  it  not  at  all  toxic. 

The  following  experiments  were  performed  in  the  study  of 
the  toxic  properties  of  potassium  sulfocyanate.8     The  details 
of  procedure  will  be  given  with  each  experiment. 
The   Effect   of   Injections   of    Various   Amounts   of   Potassium 

Sulfocyanate  into  the  Dorsal  Lymph  Sacs  of  Frogs. 

(a)  Injected  into  the  dorsal  lymph  sac  of  a  frog  weighing 
25.5  grams,  1  cc.  of  a  5  per  cent,  solution  of  potassium  sulfo- 

1  Brunton:  Pharmacology,  Therapeutics  and  Materia  Medica,  1893, 
p.  114. 

2  Kletzinski:  Heller's  Archiv.,  1833,  p.  39. 

3  Nerking:  Med.  Klinik,  191 1,  cit.  after  Diena. 

4  Lehman:  Arch.  f.  Zahnheilk.,  1911,  cit.  after  Diena. 

5  Diena:  Biochem.  Zeitschr.,  1912,  xxxix,  p.  13. 

8  The  KSCN  used  was  Kahlbaum's.  In  order  to  make  certain  that  no 
cyanide  was  present,  several  large  portions  were  tested  according  to  the 
directions  of  A.  A.  Noyes,  (Jour.  Amer.  Chem.  Soc,  xxxiv,  1912,  p.  609). 
No  trace  of  cyanide  was  found. 


75 

cyanate  (50  mg.).  Frog  had  violent  strychnine  convulsions 
with  marked  opisthotonus.  Died  in  two  minutes.  Heart 
stopped  in  systole. 

(6)  Injected  into  dorsal  lymph  sac  of  frog  (weight  27 
grams)  1  cc.  of  4  per  cent,  solution  of  potassium  sulfocyanate 
(40  mg.).  Frog  went  into  spasms;  opisthotonus;  died  in 
35  minutes.      Heart  stopped  in  systole. 

(c)  Injected  into  dorsal  lymph  sac  of  frog  (weighing  21.5 
grams)  1  cc.  of  30  per  cent,  potassium  sulfocyanate  solution 
(30  mg.).  Frog  had  spasms;  died  in  tetany  in  45  minutes. 
Heart  stopped  in  systole. 

(d)  Injected  into  dorsal  lymph  sac  of  frog  (weight  38.8 
grams)  1  cc.  of  2  per  cent,  potassium  sulfocyanate  solution. 
Frog  sickened,  but  had  no  spasms.      Died  in  41  ,  hours. 

These  experiments  were  repeated  several  times  with 
practically  the  same  results. 

(e)  Injected  into  dorsal  lymph  sac  of  frog  (weight  19.5 
grams)  7.5  mg.  potassium  sulfocyanate.  Died  in 
opisthotonus  in  fourteen  hours. 

(/)  Injected  into  dorsal  lymph  sac  of  frog  (21.5  grams)  7.5 
mg.  of  potassium  sulfocyanate.  Frog  died  in  opisthotonus  in 
sixteen  hours. 

(g)  Injected  into  dorsal  lymph  sac  of  frog  (weight  20.5 
grams)  5  mg.  of  potassium  sulfocyanate  on  April  2,  191 2. 
Frog  was  hypersensitive,  went  into  extreme  tetanic  contrac- 
tion upon  the  slightest  stimulation.  The  hypersensitiveness 
lasted  for  three  days.  On  April  9,  191 2,  frog  still  well,  has 
lost  its  irritability.     Seems  normal. 

(h)  Exact  duplication  of  experiment  g.  Frog  weighed 
18.5  grams.  Dose  was  5  mg.  of  potassium  sulfocyanate. 
Recovered  completely. 

Effect  on  Subcutaneous  Injections  of  Solutions  of  Potassium 
Sulfocyanate  on  Guinea  Pigs. 
Experiment  1. — Injected  subcutaneously  into  a  guinea  pig, 
weighing  430  gms.  500  mgs.  of  KSCN.  The  pig  died  in  one 
hour  in  marked  convulsions.  This  dose  was  also  given  hypo- 
dermically  to  another  pig,  weighing  412  gms.  This  animal  died 
in  violent  convulsions  in  two  hours  after  injection. 


76 

Experiment  2. — Into  a  guinea  pig,  weighing  350  gms.,  I  in- 
jected subcutaneously  1  cc.  of  a  20%  solution  KSCN  (200  mgs.) 
The  pig  was  found  dead  next  morning. 

Experiment  3. — Injected  into  a  guinea  pig,  weighing  385 
gms.,  1  cc.  of  a  10%  solution  KSCN  (100  mgs.)  The  animal 
expired  next  morning  in  very  violent  tetanic  convulsions. 

Experiment  4. — A  hypodermic  injection  of  1  cc.  of  a  5% 
solution  KSCN  (50  mgs.)  had  a  sickening  effect  upon  the  guinea 
pig,  the  animal  had  diarrhoea  and  did  not  eat  its  food,  After 
24  hours  it  began  to  have  convulsions  and  finally  died.  Animal 
weighed  335  grams. 

Experiment  5. — The  injection  of  25  mgs.  produced  no  visible 
effects  upon  a  guinea  pig  weighing  415  grams. 

Effect  on  the  Administration  of  Potassium  Sulfocyanate,  by  Mouth. 

Experiment  1. — A  dog,  weighing  8.1  kilos,  was  given  0.61 
gram  KSCN  in  a  meat  ball  on  each  of  three  consecutive  days. 
Calculated,  the  amount  given  is  equal  to  75  mgs.  KSCN  per 
kilo  of  dog,  or  about  1V4  grains.  After  the  second  dose,  the 
dog  did  not  partake  of  his  food,  vomited,  had  tremors  all  over 
body.  The  dog  died  in  convulsions  twenty-four  hours  after 
the  third  dose.  The  analysis  of  the  urine  showed  that  most 
of  the  sulfocyanate  was  excreted  by  the  kidneys. 

Experiment  2. — Administered  to  a  dog,  weighing  10.8  kilos, 
50  mgs.  KSCN  per  kilo  of  the  animal.  The  dog  became  de- 
pressed, vomited  and  had  diarrhoea.  The  condition  did  not 
seem  to  become  worse  after  five  findings. 

Experiment  3. — To  a  dog,  weighing  6.12  kilos,  I  gave  by 
mouth  200  mgs.  KSCN  in  a  meat  ball  on  each  of  five  consecu- 
tive days.  The  dog  suffered  from  depression,  tremors,  diar- 
rhoea, vomiting.  Recovered  upon  cessation  of  administration 
of  the  sulfocyanate. 

Experiment  4. — A  dose  of  25  mgs.  per  each  kilo  of  weight  of 
a  dog,  weighing  9.4  kilos  produced  no  toxic  effects  that  were 
discernible. 

It  may  be  mentioned  here  that  Breton,  Bruyant  and  Mezie1 
recently  reported  that  they  injected  0.03  gm.  NH4SCN  into  the 

1  M.  Breton,  L.  Bruyant  and  A.  Mczie:  Compt.  rend.  soc.  biol.,  LXXII, 
1912,  p.  400. 


77 

jugular  veins  of  dogs.     They  recovered  the  thiocyanate  in  one 
half  hour  in  the  urine.     They  do  not  refer  to  the  toxic  effects. 

The   Effects  of  Solutions  of   Potassium  Sulfocyanide  upon   //(. 
Germination  and  Growth  of  Plants. 

Raulin  found  that  in  growing  the  Aspergillus  fumigaius, 
it  was  necessary  to  add  iron  salts  to  the  spraying  water  to 
neutralize  a  certain  poison  in  the  plant  which  prevents  the 
growth  of  the  plant.  This  toxic  principle  he  found  to  be 
sulfocyanic  acid. 

Chouppe  found  saliva  fatal  to  vegetable  life.  He  de- 
termined this  fact  by  watering  seedlings  with  saliva  only. 
He  described  this  deleterious  influence  to  the  sulfocyanate 
content  of  the  saliva. 

Experiment  to  Determine  Toxic  Effects  of  Potassium  Thio- 
cyanate on  White  Lupins. 
Selected  white  lupin  beans  (Lupinus  alba)  were  soaked  in 
distilled  water  over  night.  They  were  then  carefully  planted 
in  moss  which  had  been  soaked  in  water  and  washed  clear 
of  all  visible  impurities.  When  the  roots  had  sprouted  to 
be  from  three-quarters  of  an  inch  to  an  inch  in  length  they 
were  taken  from  the  moss  and  treated  as  follows:  Jena 
glass  beakers  (400  cc.  volume)  were  carefully  washed  with 
soap  and  water  and  then  with  solutions  of  acid  and  alkali. 
They  were  then  rinsed  with  distilled  water.  Into  each  beaker 
were  placed  200  cc.  of  various  dilutions  of  potassium  sulfo- 
cyanate. The  beans  were  taken  out  from  the  moss,  were 
carefully  wiped  off  on  a  smooth  soft  cloth  and  the  shells  were 
carefully  peeled.  Upon  the  root  of  each  seedling  a  mark 
was  placed  15  mm.  from  the  tip.  India-ink  was  used  for  the 
marking.  The  bean  was  then  carefully  speared  upon  a 
sharp  glass  rod  which  was  supported  by  a  cork  plate  covering 
the  beaker.  The  rod  was  lowered  so  that  only  the  root  of 
seedling  was  in  the  liquid,  the  body  did  not  come  in  contact 
with  the  solution.  The  solutions  used  were  as  follows: 
5  per  cent.,  4  per  cent.,  3  per  cent.,  2  per  cent.,  1  per  cent., 
3/4  per  cent.,  l/i  Per  cent.,  lft  per  cent.,  2/5  per  cent.,  '  s  per 


78 

cent.,  V10  per  cent.,  V20  Per  cent->  1Im  Per  cent->  and  V100  Per 
cent.  Control  tests  were  made  with  distilled  water  and  tap 
water. 

The  following  observations  were  made  daily:  The  length 
of  the  root,  the  amount  of  chlorophyl  formation,  the  growth 
of  side  roots,  the  growth  of  tufts,  the  amount  of  mold  growth 
in  the  solutions. 

Concentrations  of  potassium  sulfocyanate  from  0.2-5.0 
per  cent,  are  completely  toxic  to  these  plants.  The  roots 
do  not  grow,  they  assume  a  white,  waxy,  oedematous  ap- 
pearance, no  side  roots  are  formed,  and  not  tufts  make  their 
appearance.  Chlorophyl  formation  though  inhibited,  is 
still  present  in  all  the  beans.  The  solution  showed  very 
heavy  growths  of  various  fungi  and  protozoa. 

The  dilute  solutions  of  the  thiocyanate  from  0.01-0.1  per 
cent,  inhibit  the  growth  of  the  roots  proportionately  to  the 
amount  of  the  noxious  substance  present.  The  roots  grow 
to  a  considerable  length  with  the  0.01  per  cent,  solution, 
side  roots  are  formed  in  various  number,  chlorophyl  formation 
is  present,  and  the  growth  of  molds  is  very  much  less.  Tuft 
growth  is  quite  marked  in  the  0.01  per  cent.,  0.02  per  cent., 
0.025  per  cent.,  0.05  per  cent,  solutions  and  very  slight  in  the 
0.1  per  cent,  solution. 

Compared  with  the  two  controls  of  distilled  water  and  tap 
water  even  the  0.0 1  per  cent,  solution  of  potassium  thio- 
cyanate is  quite  toxic;  for,  the  growth  of  the  main  root  and 
side  branches,  as  well  as  the  growth  of  tufts,  is  strongly  in- 
hibited by  the  0.0 1  per  cent,  dilution. 

Experiments  to  Determine  Toxic  Effects  of  Potassium  Thio- 
cyanate on  Timothy  Grass  Seedlings. 
Disks  of  white  blotting  paper  were  cut  two  inches  in 
diameter.  They  were  allowed  to  soak  in  distilled  water  for 
five  minutes.  Each  disk  of  paper  was  put  in  a  flat  petri 
dish  and  sprinkled  evenly  with  timothy  grass  seed.  The 
disk  was  then  covered  with  a  glass  beaker,  and  was  sprayed 
daily  with  pure  water  until  the  grass  blades  were  about  Via 
inch  in  height. 


79 

Solutions  of  potassium  sulfocyanate  were  made  of  various 
dilutions,  o.oi-io.o  per  cent.  A  separate  grass  plot  was 
used  for  each  dilution.  A  control  was  kept  with  distilled 
water.  Each  disk  was  watered  twice  daily  with  five  drops 
of  the  respective  solutions.  The  toxic  effects  of  the  sulfo- 
cyanate were  noticed  in  two  days.  The  grass  blades  did  not 
increase  in  size,  became  yellowish  green  in  color,  and  finally 
after  seven  days,  shrivelled  and  blackened.  A  profuse 
growth  of  molds  was  noticed  in  all  the  plots  which  had  been 
sprayed  with  potassium  sulfocyanate.  The  control  plot 
showed  an  exuberant  growth  of  delicate,  slender,  elongated 
blades  almost  two  inches  in  length,  without  any  mold  growth, 

Several  repetitions  were  made  of  these  experiments,  and 
almost  identical  results  were  obtained.  The  experiment 
was  then  varied.  Instead  of  watering  the  plant  daily  with 
the  sulfocyanate  solution  it  was  only  sprayed  six  times  with  five 
drops  of  the  toxic  substance.  After  the  third  day  distilled  water 
was  used.  The  objection  to  spraying  the  plant  daily  with  the 
thiocyanate  solution  is  that  as  the  water  evaporates  the  sulfo- 
cyanate remains  at  the  roots  of  the  plant  and  the  daily 
concentration  of  this  salt  becomes  steadily  greater. 

However,  even  with  the  precaution  just  now  detailed,  the 
toxic  effects  of  the  sulfocyanate  was  quite  evident.  The 
growth  was  stunted,  and  mold  growth  was  prolific  in  direct 
proportion  to  the  amount  of  potassium  sulfocyanate. 

The  Effect  of  KSCN  Solutions  on  the  Germination  of  Seeds. 

White  Lupins  that  were  allowed  to  soak  for  eighteen  hours 
in  solutions  of  greater  concentration  than  0.02%  did  not  germi- 
nate upon  being  transferred  to  beakers  containing  distilled 
water.  The  growth  of  molds  in  these  beakers  was  very  ex- 
uberant. A  0.01%  solution  of  KSCN  did  not  prevent  the 
germination  of  the  beans. 

Upon  timothy  seedlings  potassium  sulfocyanate,  even  in 
dilutions  of  0.01%,  completely  inhibits  germination. 

Bactericidal  Properties  of  Potassium  Sulfocyanate. 
Tubes  containing  gelatin  and  gelatin-peptone  media  were 
inoculated  with  bacteria.     To  each  tube  was  added  a  different 


8o 

amount  of  potassium  sulfocyanate.  Control  tubes  were 
also  kept.  The  tubes  were  allowed  to  stand  at  room  tem- 
perature for  several  days.  The  growth  in  each  tube  was 
noticed. 

It  was  observed  that  culture  media  containing  less  than  3% 
KSCN  did  not  inhibit  the  growth  of  bacteria.  A  6%,  7%,  8%, 
9%  and  10%  KSCN  gelatin  seemed  to  retard  but  did  not  com- 
pletely inhibit  the  growth  of  dust  bacteria.  The  growth  of 
molds  in  every  tube  was  very  luxuriant.  Though  the  growth 
of  the  bacteria  was  quite  marked  in  every  case,  still  the  retar- 
dation in  growth  is  directly  proportional  to  the  amount  of  sulfo- 
cyanate present. 

Effect  of  Potassium  Sulfocyanate  on  Yeast  Fermentation. 

A  5  per  cent,  glucose  solution  was  made  up  and  put  into  a 
number  of  Einhorn  saccharometers.  To  the  various 
saccharometers  I  added  various  amounts  of  potassium  sulfo- 
cyanate. A  control  tube  to  which  no  sulfocyanate  had  been 
added,  was  also  made. 

A  uniform  suspension  was  prepared  of  the  best  yeast  in 
distilled  water.  To  each  saccharometer  I  added  1  cc.  of  this 
suspension.  The  amount  of  fermentation  was  determined 
by  the  quantity  of  carbon  dioxide  produced  in  each  saccha- 
rometer. 

No  inhibition  in  the  amount  of  fermentation  was  noticed. 
In  fact,  if  anything,  the  growth  of  the  yeast  in  the  sugar  solu- 
tions seemed  to  be  stimulated  by  the  presence  of  high  amounts 
of  potassium  thiocyanate. 

Effect  of  KSCN  on  the  Souring  of  Milk. 

To  a  number  of  test  tubes  containing  milk,  I  added  varying 
amounts  of  potassium  sulfocyanate  and  a  few  drops  of  litmus 
solution.  Solutions  containing  0.5%,  1%,  2%,  3%  and  4% 
of  KSCN  prevented  the  coagulation  of  milk  even  after  four 
days.  Lower  amounts  of  sulfocyanate  in  milk  were  without 
noticeable  effect  in  this  regard. 

The  amount  of  acid  production  is  inversely  proportional  to 
the  amount  of  potassium  sulfocyanate  present.  Upon  titrating 
five  cc.  of  filtered  milk  from  each  of  six  tubes  to  which  varying 


8i 

amounts  of  KSCN  were  added,  I  found  that,  compared  with 
the  control  to  which  no  KSCN  was  added  there  was  a  marked 
reduction  in  the  quantity  of  acid  formation.  In  these  titrations 
NaOH  solution  (i  cc.  =  0.0079  Sras-  NaOH)  was  used,  phenol 
phthalein  being  the  indicator.  The  results  are  recorded  in  the 
accompanying  table. 

Amount  of  Acid  Formation  in  Milk  upon  adding  varying  amounts 

of  KSCN. 


Per  cent. 

Amt.  NaOH 

Number. 

Amt.  of  Milk. 

KSCN. 

cc. 

Control 

5  cc. 

O.O 

1.4 

I 

5  cc 

O.06 

I  .2 

II 

5  cc. 

O.  12 

I  .2 

III 

5  cc. 

O.24 

I  .  I 

IV 

5  cc. 

O.48 

O.9 

V 

5  cc. 

095 

O.8 

VI 

5  cc. 

I  .90 

0.3 

(Several  experiments  along  these  lines  are  in  progress,  and 
will  be  described  later.  When  this  dissertation  went  to  press, 
the  available  data  of  the  later  experiments  were  not  sufficiently 
complete  for  insertion.) 

conclusions. 

1.  Potassium  sulfocyanate  is  toxic  to  both  plants  and 
animals.  Its  toxicity  is  so  marked  that  indiscriminate  dispen- 
sation of  the  substance  to  patients  is  dangerous. 

2.  The  growth  of  molds  is  enhanced  by  potassium  sulfo- 
cyanate. 

3.  Small  biological  quantities  of  KSCN  have  no  inhibiting 
influence  on  bacteria. 

4.  Yeast  fermentation  is  uninhibited  or  stimulated  by  potas- 
sium sulfocyanate. 

5.  The  souring  of  milk  is  inhibited  by  large  (relatively) 
amounts  of  the  thiocyanate. 


ADDENDUM. 
Bibliography  not  specifically  alluded  to  in  the  body  of  this  disserta 
tion. 

i.  Archetti,  Chem.  Centralblatt,  1904,  i,  pp.  318,  371  and  796. 

2.  Acr'ee  and  Hickins,  Dental  Review,  xxii,  p.  219. 

3.  Bujuid,  Virchow's  Arch.,  xci,  p.  190. 

4.  Benson,  Jahresb.  d.  fort.  d.  Chem.,  1852,  p.  439. 

5.  Berkson,  The  Odontologist,  191 1,  vii,  p.  15. 

6.  Beach,  Dental  Cosmos,  1908,  1,  p.  469. 

7.  Bentley  and  Le  Roy,  N.  Y.  Med.  Jour.,  1908,  Ixxxix,  p.  210. 

8.  Buttazzoni,  Stomatol.  Milano,  1902,  i,  p.  764. 

9.  Bergman,  Therapie  der  Gegenwort,  1903,  p.  200. 

10.  Claisse  and  Dupre,  Compt.  rend.  soc.  de  Biol.,  1894,  vi,  p.  55. 

11.  Chittenden  and  Richards,  Am.  J.  Phys.,  1898,  i,  p.  461. 

12.  Carlson  and  McLean,  Am.  Jour.  Physiol.,  1908,  xx,  p.  457. 

13.  Duclaux,  Le  Microbe  et  la  Maladie,  Vol.  iv. 

14.  Doubleday,  Dental  Cosmos,  1909,  li,  p.  412. 

15.  G.  Deniges,  Jour.  d.  Pharm.,  1898,  xxv,  p.  88. 

16.  Edinger  and  Clemens,  Z.  /.  klin.  Med.,  1906,  lix,  p.  223. 

17.  Goadley,  Brit.  Med.  Jour.,  1910,  p.  770. 

18.  Gmelin,  Handb.  d.  Chem. 

19.  Goodman,  Proc.  Path.  Soc.  Philadel.,  1909,  xxii,  p.  9. 

20.  Gautrand,  Du  chimisme  salivaire,  Lyon,  1895. 

21.  Grawitz  and  Steffen,  Berl.  Klin.  Woch.,  1894,  xxxi,  p.  419. 

22.  Hofbauer,  Arch.  f.  d.  ges.  Physiol.,  1896,  lxv,  p.  503. 

23.  Jappeli,  Zeit.  f.  Biologie,  1908,  li,  p.  127. 

24.  Jamison,  Ann.  d.  Chem.  u.  Pharm.,  lviii,  p.  264. 

25.  Klason,  Jour,  prakt.  Chem.,  xxxv,  p.  407. 

26.  Liebig,  Ann.  d.  Chem.  u.  Pharm.,   lxi,  p.  126. 

27.  Lidow,  Jour.  Russ.  Chem.  Soc,  xvi,  p.  271. 

28.  Langley,  Trans.  Roy.  Phil.  Soc,  1880,  clxxx,  p.  109. 

29.  Marcantonio,  Rejorma  Med.  Napoli,  1897,  xiii,  p.  254. 

30.  Mislawsksky  and  Smirnow,  Arch.  f.  Physiol.  Leipzig,  1896,  p.  93. 

31.  Mensel,  Die  Quellkraft  d.  Rhodanate,  1886. 

32.  Nencki,  Ber.  deut.  chem.  Ges.,  xxvii,  p.  1318. 

33.  Nollner,  Jahresb.  d.  Fortsch.  d.  Chem.,  1856,  p.  443. 

34.  Neumeister,  Physiol,  d.  Menschen,  p.  844.  " 

35.  Ostwald,  Jour,  prakt.  Chem.,  xxxii,  p.  305. 

36.  Pribram,  Jahr.  d.  Phys.  u.  Path.,  1868,  i,  p.  148. 

37.  Pettenkofer,  Chem.  Centralbl.,  1846,  p.  231. 

38.  Pozzi,  Chem.  Centralbl.,  1904,  ii,  p.  331. 

39.  Radiszewski,  Ann.  d.  Chem.  u.  Pharm. 

40.  Sticker,  Munch,  med.  Woch.,  1896,  xliii,  p.  1041. 


«3 

4i.  Schoumow-Simanowsky,    Arch.   j.    exp.    path.    u.    Pharm.,    1891, 
xxxiv,  p.  332. 

42.  Sertoli,  Virchow's  Hirsch  Jahresb.,  1869,  >i  P-  '04. 

43.  Schiff,  Ann.  d.  Chem.  u.  Pharm.,  cvi,  p.  116. 

44.  Tozner,  Arch.  int.  Physiol.,  1905,  ii,  p.  153. 

45.  Thiel,  Ber.  deut.  chem.  Ges.,  1902,  xxxv,  p.  2766. 

46.  Triolo,  Jahresber.  d.  Tierchem.,  1897,  xxvii,  p.  813. 

47.  Ville  and  Mestrezat,  Compt.  rend.  Soc.  d.  Biol.,  65,  66. 

48.  Volckel,  Jahr.  d.  Fort.  d.  Chem.,  1853,  p.  406. 

49.  Von  Beustl,  Brit.  Dent.  Jour.,  1911,  xxxii,  p.  65. 

50.  Van  Setten,  Muller  Arch.  d.  Anat.,  1838,  p.  164. 

51.  Volhard,  Ann.  d.  chem.  u.  Pharm.,  cxc,  p.  24. 

52.  Warner,  Chem.  Centralbl.,  1903,  i,  p.  985. 

53.  Weber,  Jahresb.  d.  Tierchem.,  1892,  xxii,  p.  245. 

54.  Zeise,  Ann.  d.  Chem.  u.  Pharm.,  xlvii,  p.  36. 

55.  Zimmerman,  Ann.  d.  Chem.  u.  Pharm.,  excix,  p.  1. 


BIOGRAPHICAL. 
Max  Kahn  was  born  on  March  17,  1887.  In  1905  he  ob- 
tained a  scholarship  in  Cornell  University  which  he  held 
until  1 9 10,  when  he  was  granted  the  degree  of  Doctor  of 
Medicine  from  that  institution.  During  the  years  1910-1912, 
he  pursued  graduate  work  in  Columbia  University.  In 
June.  191 1,  he  received  the  degree  of  Master  of  Arts,  his 
major  subject  being  Organic  Chemistry,  his  thesis  being 
entitled  'The  Chemistry  of  the  Xaphthodiazines."  He  held 
the  University  Scholarship  in  Columbia  University  for  the 
year  1911-1912.  during  which  time  he  pursued  studies  in 
Biological  Chemistry  for  the  degree  of  Doctor  of  Philosophy. 
He  has  lately  been  appointed  Director  of  the  Chemical  and 
Physiological  Laboratories  of  Beth  Israel  Hospital,  New  York 
City. 


PUBLICATIONS. 

1.  Shakespeare's    knowledge    of    medicine.     New     York    Medical 
Journal,  1910,  xcii,  p.  957. 

2.  Moliere  and   the  physician.     Johns  Hopkins  Hospital  Bulletin, 
1911,  xxii,  p.  344. 

3.  The  importance  of  the  colloidal  nitrogen  in  the  urine,  ia  the 
diagnosis  of  cancer.     American  Journal  of  Gastroenterology,  191 1,  i,  p.  11. 

4.  Ueber  den  Wert  des  colloidalen  Stickstoffs  im  Urin  bei  der  Krebs- 
diagnose.     Archiv.  fur  Verdauungs  Krankheiten,  1911,  xvii,  p.  557. 

5.  On  the  absorption  and  distribution  of  aluminium  from  aluminized 
food.     Biochemical  Bulletin,  1911,  i,  p.  235. 

6.  The  colloid  nitrogen  content  of  normal  and  cancerous  dog's  urine. 
(In  press.) 

7.  The  chemistry  of  renal  and  cystic  calculi.     (In  press.) 

8.  History  of  the  lithotomy  operation.     (In  press.) 

9.  Rambam  the  physician.     (In  press.) 

10.  Therapeutic  superstitions  and  vulgar  specifics.     Medical  Record. 
(In  press.) 


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